ST receptor binding compounds and methods of using the same

ABSTRACT

Conjugated compounds which comprises an ST receptor binding moiety and a radiostable active moiety are disclosed. Pharmaceutical compositions comprising a pharmaceutically acceptable carrier or diluent, and a conjugated compound which comprises an ST receptor binding moiety and a radiostable active moiety or an ST receptor binding moiety and a radioactive active moiety are disclosed. Methods of treating an individual suspected of suffering from metastasized colorectal cancer comprising the steps of administering to said individual a pharmaceutical composition comprising a pharmaceutically acceptable carrier or diluent, and a therapeutically effective amount of a conjugated compound which comprises an ST receptor binding moiety and a radiostable active moiety or an ST receptor binding moiety and a radiostable active moiety are disclosed. Methods of radioimaging metastasized colorectal cancer cells comprising the steps of first administering to an individual suspected of having metastasized colorectal cancer cells, a pharmaceutical composition that comprises a pharmaceutically acceptable carrier or diluent, and conjugated compound that comprises an ST receptor binding moiety and a radioactive active moiety wherein the conjugated compound is present in an amount effective for diagnostic use in humans suffering from colorectal cancer and then detecting the localization and accumulation of radioactivity in the individual&#39;s body are disclosed.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.08/468,449 filed Jun. 6, 1995, which is a continuation of and claimspriority to U.S. application Ser. No. 08/141,892, filed Oct. 26, 1993,now U.S. Pat. No. 5,518,888, the entire contents of which areincorporated herein by reference.

ACKNOWLEDGEMENT OF GOVERNMENT RIGHTS

This invention was made with Government support under grant numberDK43805-01A2 awarded by the National Institutes of Health. TheGovernment has certain rights in this invention.

FIELD OF THE INVENTION

The present invention relates to compounds which comprise a receptorligand moiety conjugated to an active agent. More particularly, thepresent invention relates to compounds which comprise a moiety thatbinds to the ST receptor conjugated to a therapeutic or imaging moiety.

BACKGROUND OF THE INVENTION

Colorectal cancer is the third most common neoplasm worldwide and thesecond most common in the United States, representing about 15% of thenewly diagnosed cases of cancer in the United States. The largeintestine or large bowel is the third leading site for the developmentof new cancer and is diagnosed in about 150,000 patients each year.Colorectal cancer is the second leading cause of cancer-related deathsand is responsible for about 12% of cancer deaths in the United States.The mortality rate of newly diagnosed large bowel cancer approaches 50%and there has been little improvement over the past 40 years. Most ofthis mortality reflects local, regional and distant metastases. Aboutthirty percent of patients with colorectal cancer have unresectabledisease at presentation and about 40% develop metastases during thecourse of their disease. Distant metastatic disease is seen in liver(about 12%), lung (about 3%), bone (about 0.9%), brain (about 0.7%),nodes (about 4%), and peritoneum (about 2%) at the time of initialdiagnosis. In 1987, the large bowel cancers found regionally or atdistant sites at the time of diagnosis were about 26% and about 18%,respectively.

Surgery is the mainstay of treatment for colorectal cancer butrecurrence is frequent. Colorectal cancer has proven resistant tochemotherapy, although limited success has been achieved using acombination of 5-fluorouracil and levamisole. Surgery has had thelargest impact on survival and, in some patients with limited disease,achieves a cure. However, surgery removes bulk tumor, leaving behindmicroscopic residual disease which ultimately results in recrudescence.Overall recurrence rates for colonic tumors are about 33% and for rectalcancer about 42%. Of these recurrences, about 9% are local, about 13%are systemic metastatic disease, and the remaining 88% are a combinationof local and systemic disease. Fifty percent of patients with recurrentcolorectal cancer have hepatic metastases.

Early detection of primary, metastatic, and recurrent disease cansignificantly impact the prognosis of individuals suffering fromcolorectal cancer. Large bowel cancer diagnosed at an early stage has asignificantly better outcome than that diagnosed at more advancedstages. The 5 year relative survival rates for patients with regional ordistant metastases are 48% and 5%, compared with 90% and 77% for diseasewhich is in situ or local, respectively, at the time of diagnosis.Similarly, diagnosis of metastatic or recurrent disease earlierpotentially carries with it a better prognosis.

Although current radiotherapeutic agents, chemotherapeutic agents andbiological toxins are potent cytotoxins, they do not discriminatebetween normal and malignant cells, producing adverse effects anddose-limiting toxicities. Over the past decade, a novel approach hasbeen employed to more specifically target agents to tumor cells,involving the conjugation of an active agent to molecules which bindspreferentially to antigens that exist predominantly on tumor cells.These conjugates can be administered systemically and specifically bindto the targeted tumor cells. Theoretically, targeting permits uptake bycells of cytotoxic agents at concentrations which do not produce serioustoxicities in normal tissues. Also, selective binding to targeted tumorcells facilitates detection of occult tumor and is therefore useful indesigning imaging agents. Molecular targeting predominantly has employedmonoclonal antibodies generated to antigens selectively expressed ontumor cells.

Immunoscintigraphy using monoclonal antibodies directed attumor-specific markers has been employed to diagnose colorectal cancer.Monoclonal antibodies against carcinoembryonic antigen (CEA) labeledwith ⁹⁹Technetium identified 94% of patients with recurrent tumors.Similarly, ¹¹¹Indium-labeled anti-CEA monoclonal antibodies successfullydiagnosed 85% of patients with recurrent colorectal carcinoma who werenot diagnosed by conventional techniques. ¹²⁵Iodine-labeled antibodieshave been effective in localizing more than 80% of thepathologically-confirmed recurrences by intraoperative gamma probescanning.

Monoclonal antibodies have also been employed to target specifictherapeutic agents in colorectal cancer. Preclinical studiesdemonstrated that anti-CEA antibodies labeled with ⁹⁰Yttrium inhibitedhuman colon carcinoma xenografts in nude mice. Antibodies generated tocolorectal cancer cells and coupled to mitomycin C or neocarzinostatindemonstrated an anti-tumor effect on human colon cancer xenografts innude mice and 3 patients with colon cancer. Similar results in animalswere obtained with monoclonal antibodies conjugated to ricin toxin Achain. Due to the sensitivity, specificity, and adverse-effect profileof monoclonal antibodies, the results obtained using monoclonalantibody-based therapeutics have shown them to be less than idealtargeting tools. Although monoclonal antibodies have been generated toantigens selectively expressed on tumors, no truly cancer-specificantibody has been identified. Most antigens expressed on neoplasticcells appear to be quantitatively increased in these compared to normalcells but the antigens are nonetheless often present in normal cells.Thus, antibodies to such determinants can react with non-neoplastictissues, resulting in significant toxicities. Also, antibodies arerelatively large molecules and consequently, often evoke an immuneresponse in patients. These immune responses can result in significanttoxicities in patients upon re-exposure to the targeting agents and canprevent targeting by the monoclonal due to immune complex formation withdegradation and excretion. Finally, binding of antibodies to tumor cellsmay be low and targeted agents may be delivered to cells in quantitiesinsufficient to achieve detection or cytotoxicity.

There remains a need for compositions which can specifically targetmetastasized colorectal cancer cells. There is a need for imaging agentswhich can specifically bind to metastasized colorectal cancer cells.There is a need for improved methods of imaging metastasized colorectalcancer cells. There is a need for therapeutic agents which canspecifically bind to metastasized colorectal cancer cells. There is aneed for improved methods of treating individuals who are suspected ofsuffering from colorectal cancer cells, especially individuals who aresuspected of suffering from metastasis of colorectal cancer cells.

SUMMARY OF THE INVENTION

The present invention relates to conjugated compounds which comprises anST receptor binding moiety and a radiostable active moiety.

The present invention relates to a pharmaceutical composition comprisinga pharmaceutically acceptable carrier or diluent, and a conjugatedcompound which comprises an ST receptor binding moiety and a radiostableactive moiety.

The present invention relates to a method of treating an individualsuspected of suffering from metastasized colorectal cancer comprisingthe steps of administering to said individual a pharmaceuticalcomposition comprising a pharmaceutically acceptable carrier or diluent,and a therapeutically effective amount of a conjugated compound whichcomprises an ST receptor binding moiety and a radiostable active moiety.

The present invention relates to a pharmaceutical composition comprisinga pharmaceutically acceptable carrier or diluent, and conjugatedcompound that comprises an ST receptor binding moiety and a radioactiveactive moiety wherein the conjugated compound is present in an amounteffective for therapeutic or diagnostic use in humans suffering fromcolorectal cancer.

The present invention relates to a method of radioimaging metastasizedcolorectal cancer cells comprising the steps of first administering toan individual suspected of having metastasized colorectal cancer cells,a pharmaceutical composition that comprises a pharmaceuticallyacceptable carrier or diluent, and conjugated compound that comprises anST receptor binding moiety and a radioactive active moiety wherein theconjugated compound is present in an amount effective for diagnostic usein humans suffering from colorectal cancer and then detecting thelocalization and accumulation of radioactivity in the individual's body.

The present invention relates to a method of treating an individualsuspected of suffering from metastasized colorectal cancer comprisingthe steps of administering to said individual a pharmaceuticalcomposition comprising a pharmaceutically acceptable carrier or diluent,and a therapeutically effective amount of a conjugated compound whichcomprises an ST receptor binding moiety and a radioactive active moiety.

DESCRIPTION OF PREFERRED EMBODIMENTS OF THE INVENTION

As used herein, the terms “ST” and “native ST” are used interchangeablyand are meant to refer to heat-stable toxin (ST) which is a peptideproduced by E. coli, as well as other organisms. STs are naturallyoccurring peptides which 1) are naturally produced by organisms, 2)which bind to the ST receptor and 3) which activate the signal cascadethat mediates ST-induced diarrhea.

As used herein, the term “ST receptor” is meant to refer to thereceptors found on colorectal cells, including local and metastasizedcolorectal cancer cells, which bind to ST. In normal individuals, STreceptors are found exclusively in cells of intestine, in particular incells in the duodenum, small intestine (jejunum and ileum), the largeintestine, colon (cecum, ascending colon, transverse colon, descendingcolon and sigmoid colon) and rectum.

As used herein, the term “ST receptor ligand” is meant to refer tocompounds which specifically bind to the ST receptor. ST is an STreceptor ligand. An ST receptor ligand may be a peptide or anon-peptide.

As used herein, the term “ST receptor binding peptide” is meant to referto ST receptor ligands that are peptides.

As used herein, the term “ST peptides” is meant to refer to ST receptorbinding peptides selected from the group consisting of SEQ ID NO:2, SEQID NO:3, SEQ ID NOS:5-54 and fragments and derivatives thereof.

As used herein, the term “fragment” is meant to refer to peptide a)which has an amino acid sequence identical to a portion of an STreceptor binding peptide and b) which is capable of binding to the STreceptor.

As used herein, the term “derivative” is meant to refer to a peptide a)which has an amino acid sequence substantially identical to at least aportion of an ST receptor binding peptide and b) which is capable ofbinding to the ST receptor.

As used herein, the term “substantially identical” is meant to refer toan amino acid sequence that is the same as the amino acid sequence of anST peptide except some of the residues are deleted or substituted withconservative amino acids or additional amino acids are inserted.

As used herein, the term “active agent” is meant to refer to compoundsthat are therapeutic agents or imaging agents.

As used herein, the term “radiostable” is meant to refer to compoundswhich do not undergo radioactive decay; i.e. compounds which are notradioactive.

As used herein, the term “therapeutic agent” is meant to refer tochemotherapeutics, toxins, radiotherapeutics, targeting agents orradiosensitizing agents.

As used herein, the term “chemotherapeutic” is meant to refer tocompounds that, when contacted with and/or incorporated into a cell,produce an effect on the cell including causing the death of the cell,inhibiting cell division or inducing differentiation.

As used herein, the term “toxin” is meant to refer to compounds that,when contacted with and/or incorporated into a cell, produce the deathof the cell.

As used herein, the term “radiotherapeutic” is meant to refer toradionuclides which when contacted with and/or incorporated into a cell,produce the death of the cell.

As used herein, the term “targeting agent” is meant to refer compoundswhich can be bound by and or react with other compounds. Targetingagents may be used to deliver chemotherapeutics, toxins, enzymes,radiotherapeutics, antibodies or imaging agents to cells that havetargeting agents associated with them and/or to convert or otherwisetransform or enhance coadministered active agents. A targeting agent mayinclude a moiety that constitutes a first agent that is localized to thecell which when contacted with a second agent either is converted to athird agent which has a desired activity or causes the conversion of thesecond agent into an agent with a desired activity. The result is thelocalized agent facilitates exposure of an agent with a desired activityto the metastasized cell.

As used herein, the term “radiosensitizing agent” is meant to refer toagents which increase the susceptibility of cells to the damagingeffects of ionizing radiation. A radiosensitizing agent permits lowerdoses of radiation to be administered and still provide atherapeutically effective dose.

As used herein, the term “imaging agent” is meant to refer to compoundswhich can be detected.

As used herein, the term “ST receptor binding moiety” is meant to referto the portion of a conjugated compound that constitutes an ST receptorligand.

As used herein, the term “active moiety” is meant to refer to theportion of a conjugated compound that constitutes an active agent.

As used herein, the terms “conjugated compound” and “conjugatedcomposition” are used interchangeably and meant to refer to a compoundwhich comprises an ST receptor binding moiety and an active moiety andwhich is capable of binding to the ST receptor. Conjugated compoundsaccording to the present invention comprise a portion which constitutesan ST receptor ligand and a portion which constitutes an active agent.Thus, conjugated compounds according to the present invention arecapable of specifically binding to the ST receptor and include a portionwhich is a therapeutic agent or imaging agent. Conjugated compositionsmay comprise crosslinkers and/or molecules that serve as spacers betweenthe moieties.

As used herein, the terms “crosslinker”, “crosslinking agent”,“conjugating agent”, “coupling agent”, “condensation reagent” and“bifunctional crosslinker” are used interchangeably and are meant torefer to molecular groups which are used to attach the ST receptorligand and the active agent to thus form the conjugated compound.

As used herein, the term “colorectal cancer” is meant to include thewell-accepted medical definition that defines colorectal cancer as amedical condition characterized by cancer of cells of the intestinaltract below the small intestine (i.e. the large intestine (colon),including the cecum, ascending colon, transverse colon, descendingcolon, and sigmoid colon, and rectum). Additionally, as used herein, theterm “colorectal cancer” is meant to further include medical conditionswhich are characterized by cancer of cells of the duodenum and smallintestine (jejunum and ileum). The definition of colorectal cancer usedherein is more expansive than the common medical definition but isprovided as such since the cells of the duodenum and small intestinealso contain ST receptors and are therefore amenable to the methods ofthe present invention using the compounds of the present invention.

As used herein, the term “metastasis” is meant to refer to the processin which cancer cells originating in one organ or part of the bodyrelocate to another part of the body and continue to replicate.Metastasized cells subsequently form tumors which may furthermetastasize. Metastasis thus refers to the spread of cancer from thepart of the body where it originally occurs to other parts of the body.The present invention relates to methods of delivering active agents tometastasized colorectal cancer cells.

As used herein, the term “metastasized colorectal cancer cells” is meantto refer to colorectal cancer cells which have metastasized; colorectalcancer cells localized in a part of the body other than the duodenum,small intestine (jejunum and ileum), large intestine (colon), includingthe cecum, ascending colon, transverse colon, descending colon, andsigmoid colon, and rectum.

ST, which is produced by E. coli, as well as other organisms, isresponsible for endemic diarrhea in developing countries and travelersdiarrhea. ST induces intestinal secretion by binding to specificreceptors, ST receptors, in the apical brush border membranes of themucosal cells lining the intestinal tract. Binding of ST to ST receptorsis non-covalent and occurs in a concentration-dependent and saturablefashion. Once bound, ST-ST receptor complexes appear to be internalizedby intestinal cells, i.e. transported from the surface into the interiorof the cell. Binding of ST to ST receptors triggers a cascade ofbiochemical reactions in the apical membrane of these cells resulting inthe production of a signal which induces intestinal cells to secretefluids and electrolytes, resulting in diarrhea.

ST receptors are unique in that they are only localized in the apicalbrush border membranes of the cells lining the intestinal tract. Indeed,they are not found in any other cell type in placental mammals. Inaddition, ST receptors are almost exclusively localized to the apicalmembranes, with little being found in the basolateral membranes on thesides of intestinal cells.

Mucosal cells lining the intestine are joined together by tightjunctions which form a barrier against the passage of intestinalcontents into the blood stream and components of the blood stream intothe intestinal lumen. Therefore, the apical location of ST receptorsisolates these receptors from the circulatory system so that they may beconsidered to exist separate from the rest of the body; essentially the“outside” of the body. Therefore, the rest of the body is considered“outside” the intestinal tract. Compositions administered “outside” theintestinal tract are maintained apart and segregated from the only cellswhich normally express ST receptors.

In individuals suffering from colorectal cancer, the cancer cells areoften derived from cells that produce and display the ST receptor andthese cancer cells continue to produce and display the ST receptor ontheir cell surfaces. Indeed, T84 cells, which are human colonicadenocarcinoma cells isolated from lung metastases, express ST receptorson their cell surface. Similarly, HT29glu-cells, which are human colonicadenocarcinoma cells, express receptors for ST. Thus, in individualssuffering from colorectal cancer, some metastasized intestinal cancercells express ST receptors.

An effort was undertaken to determine the proportion of colorectaltumors which have the ST receptor. Sixteen colorectal cancer tumors,including ten local colorectal tumors and six metastasized tumors (3liver, 1 lung, 1 lymphnode, 1 peritoneum), were tested and eachpossessed ST receptors. In each case, the affinity and density ofreceptors was amenable for targeting. That is, the cells possessed atleast 10⁴-10⁶ receptors per cell and demonstrated an affinity of 10⁴ orbetter (that is preferably between 10⁻⁸ to 10⁻⁹ or less; the lowernumber indicating a tighter bond, thus a higher affinity). Normal liver,lymphnode, peritoneum and lung cells were found not to possess STreceptors.

When such cancer cells metastasize, the metastasized cancer cellscontinue to produce and display the ST receptor. The expression of STreceptors on the surfaces of metastatic tumors provides a target forselective binding of conjugated compositions. ST receptors permit theabsolutely specific targeting of therapeutic and diagnostic agents thatare conjugated to ST receptor ligands to metastatic colorectal cancercells.

The conjugated compositions of the present invention are useful fortargeting cells that line the inner intestine wall including thosecancer cells derived from such cells, particularly metastasized cancercells derived from such cells. The conjugated compositions of thepresent invention which are administered outside the intestinal tractsuch as those administered in the circulatory system will remainsegregated from the cells that line the intestinal tract and will bindonly to cells outside the intestinal tract which are derived from theintestinal tract such as metastasized colorectal cells. The conjugatedcompositions will not bind to non-colorectal derived cells. Thus, theactive moieties of conjugated compositions administered outside theintestinal tract are delivered to cells which are derived from theintestinal tract such as metastasized colorectal cells but will not bedelivered to any other cells.

Therapeutic and diagnostic pharmaceutical compositions of the presentinvention include conjugated compounds specifically targeted tometastatic disease. These conjugated compounds include ST receptorbinding moieties which do not bind to cells of normal tissue in the bodyexcept cells of the intestinal tract since the cells of other tissues donot possess ST receptors. Unlike normal colorectal cells and localizedcolorectal cancer cells, metastasized colorectal cancer cells areaccessible to substances administered outside the intestinal tract, forexample administered in the circulatory system. The only ST receptors innormal tissue exist in the apical membranes of intestinal mucosa cellsand these receptors are effectively isolated from the targeted cancerchemotherapeutics and imaging agents administered outside the intestinaltract by the intestinal mucosa barrier. Thus, metastasized colorectalcells may be targeted by conjugated compounds of the present inventionby introducing such compounds outside the intestinal tract such as forexample by administering pharmaceutical compositions that compriseconjugated compounds into the circulatory system.

One having ordinary skill in the art can readily identify individualssuspected of suffering from colorectal cancer and metastasizedcolorectal cells. In those individuals diagnosed with colorectal cancer,it is standard therapy to suspect metastasis and aggressively attempt toeradicate metastasized cells. The present invention providespharmaceutical compositions and methods for imaging and thereby willmore definitively diagnose metastasis. Further, the present inventionprovides pharmaceutical compositions comprising therapeutic agents andmethods for specifically targeting and eliminating metastasizedcolorectal cancer cells. Further, the present invention providespharmaceutical compositions that comprise therapeutics and methods forspecifically eliminating colorectal cancer cells.

The pharmaceutical compositions which comprise conjugated compositionsof the present invention may be used to diagnose or treat individualssuffering from localized colorectal tumors, that is primary ornon-metastatic colorectal tumors if these have penetrated the basementmembrane underlying the mucosa into the submucosa where there isabundant blood supply to which they have access. Penetration into thesubmucosa circumvents the mucosal barrier resulting in the ability ofconjugated compositions introduced into the circulatory system tointeract with these tumors.

The present invention relies upon the use of an ST receptor bindingmoiety in a conjugated composition. The ST receptor binding moiety isessentially a portion of the conjugated composition which acts as aligand to the ST receptor and thus specifically binds to thesereceptors. The conjugated composition also includes an active moietywhich is associated with the ST receptor binding moiety; the activemoiety being an active agent which is either useful to image, target,neutralize or kill the cell.

According to the present invention, the ST receptor binding moiety isthe ST receptor ligand portion of a conjugated composition. In someembodiments, the ST receptor ligand may be native ST.

Native ST has been isolated from a variety of organisms including E.coli, Yersinia, Enterobacter, and others. In nature, the toxins aregenerally encoded on a plasmid which can “jump” between differentspecies. Several different toxins have been reported to occur indifferent species. These toxins all possess significant sequencehomology, they all bind to ST receptors and they all activate guanylatecyclase, producing diarrhea.

ST has been both cloned and synthesized by chemical techniques. Thecloned or synthetic molecules exhibit binding characteristics which aresimilar to native ST. Native ST isolated from E. coli is 18 or 19 aminoacids in length. The smallest “fragment’ of ST which retains activity isthe 13 amino acid core peptide extending toward the carboxy terminalfrom cysteine 6 to cysteine 18 (of the 19 amino acid form) Analogues ofST have been generated by cloning and by chemical techniques. Smallpeptide fragments of the native ST structure which include thestructural determinant that confers binding activity may be constructed.Once a structure is identified which binds to ST receptors, non-peptideanalogues mimicking that structure in space are designed.

SEQ ID NO:1 discloses a nucleotide sequence which encodes 19 amino acidST, designated ST Ia, reported by So and McCarthy (1980) Proc. Natl.Acad. Sci. USA 77:4011, which is incorporated herein by reference.

The amino acid sequence of ST Ia is disclosed in SEQ ID NO:2.

SEQ ID NO:3 discloses the amino acid sequence of an 18 amino acidpeptide which exhibits ST activity, designated ST I*, reported by Chanand Giannella (1981) J. Biol. Chem. 256:7744, which is incorporatedherein by reference.

SEQ ID NO:4 discloses a nucleotide sequence which encodes 19 amino acidST, designated ST Ib, reported by Mosely et al. (1983) Infect. Immun.39:1167, which is incorporated herein by reference.

The amino acid sequence of ST Ib is disclosed in SEQ ID NO:5.

A 15 amino acid peptide called guanylin which has about 50% sequencehomology to ST has been identified in mammalian intestine (Currie, M. G.et al. (1992) Proc. Natl. Acad. Sci. USA 89:947-951, which isincorporated herein by reference). Guanylin binds to ST receptors andactivates guanylate cyclase at a level of about 10- to 100-fold lessthan native ST. Guanylin may not exist as a 15 amino acid peptide in theintestine but rather as part of a larger protein in that organ. Theamino acid sequence of guanylin from rodent is disclosed as SEQ ID NO:6.

SEQ ID NO:7 is an 18 amino acid fragment of SEQ ID NO:2. SEQ ID NO:8 isa 17 amino acid fragment of SEQ ID NO:2. SEQ ID NO:9 is a 16 amino acidfragment of SEQ ID NO:2. SEQ ID NO:10 is a 15 amino acid fragment of SEQID NO:2. SEQ ID NO:11 is a 14 amino acid fragment of SEQ ID NO:2. SEQ IDNO:12 is a 13 amino acid fragment of SEQ ID NO:2. SEQ ID NO:13 is an 18amino acid fragment of SEQ ID NO:2. SEQ ID NO:14 is a 17 amino acidfragment of SEQ ID NO:2. SEQ ID NO:15 is a 16 amino acid fragment of SEQID NO:2. SEQ ID NO:16 is a 15 amino acid fragment of SEQ ID NO:2. SEQ IDNO:17 is a 14 amino acid fragment of SEQ ID NO:2.

SEQ ID NO:18 is a 17 amino acid fragment of SEQ ID NO:3. SEQ ID NO:19 isa 16 amino acid fragment of SEQ ID NO:3. SEQ ID NO:20 is a 15 amino acidfragment of SEQ ID NO:3. SEQ ID NO:21 is a 14 amino acid fragment of SEQID NO:3. SEQ ID NO:22 is a 13 amino acid fragment of SEQ ID NO:3. SEQ IDNO:23 is a 17 amino acid fragment of SEQ ID NO:3. SEQ ID NO:24 is a 16amino acid fragment of SEQ ID NO:3. SEQ ID NO:25 is a 15 amino acidfragment of SEQ ID NO:3. SEQ ID NO:26 is a 14 amino acid fragment of SEQID NO:3.

SEQ ID NO:27 is an 18 amino acid fragment of SEQ ID NO:5. SEQ ID NO:28is a 17 amino acid fragment of SEQ ID NO:5. SEQ ID NO:29 is a 16 aminoacid fragment of SEQ ID NO:5. SEQ ID NO:30 is a 15 amino acid fragmentof SEQ ID NO:5. SEQ ID NO:31 is a 14 amino acid fragment of SEQ ID NO:5.SEQ ID NO:32 is a 13 amino acid fragment of SEQ ID NO:5. SEQ ID NO:33 isan 18 amino acid fragment of SEQ ID NO:5. SEQ ID NO:34 is a 17 aminoacid fragment of SEQ ID NO:5. SEQ ID NO:35 is a 16 amino acid fragmentof SEQ ID NO:5. SEQ ID NO:36 is a 15 amino acid fragment of SEQ ID NO:5.SEQ ID NO:37 is a 14 amino acid fragment of SEQ ID NO:5. SEQ ID NO:27,SEQ ID NO:31, SEQ ID NO:36 AND SEQ ID NO:37 are disclosed in Yoshimura,S., et al. (1985) FEBS Lett. 181:138, which is incorporated herein byreference.

SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, which are derivatives ofSEQ ID NO:3, are disclosed in Waldman, S. A. and O'Hanley, P. (1989)Infect. Immun. 57:2420, which is incorporated herein by reference. SEQID NO:41, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44 and SEQ ID NO:45,which are a derivatives of SEQ ID NO:3, are disclosed in Yoshimura, S.,et al. (1985) FEBS Lett. 181:138, which is incorporated herein byreference.

SEQ ID NO:46 is a 25 amino acid peptide derived from Y. enterocoliticawhich binds to the ST receptor.

SEQ ID NO:47 is a 16 amino acid peptide derived from V. cholerae whichbinds to the ST receptor. SEQ ID NO:47 is reported in Shimonishi, Y., etal. FEBS Lett. 215:165, which is incorporated herein by reference.

SEQ ID NO:48 is an 18 amino acid peptide derived from Y. enterocoliticawhich binds to the ST receptor. SEQ ID NO:48 is reported in Okamoto, K.,et al. Infec. Immun. 55:2121, which is incorporated herein by reference.

SEQ ID NO:49, is a derivative of SEQ ID NO:5.

SEQ ID NO:50, SEQ ID NO:51, SEQ ID NO:52 and SEQ ID NO:53 arederivatives.

SEQ ID NO:54 is the amino acid sequence of guanylin from human.

In some preferred embodiments, conjugated compounds comprise ST receptorbinding moieties that comprise amino acid sequences selected from thegroup consisting of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 andfragments and derivatives thereof.

Those having ordinary skill in the art can readily design and producederivatives having substantially identical amino acid sequences of STpeptides with deletions and/or insertions and/or conservativesubstitutions of amino acids. For example, following what are referredto as Dayhof's rules for amino acid substitution (Dayhof, M. D. (1978)Nat. Biomed. Res. Found., Washington, D.C. Vol. 5, supp. 3), amino acidresidues in a peptide sequence may be substituted with comparable aminoacid residues. Such substitutions are well-known and are based the uponcharge and structural characteristics of each amino acid. Derivativesinclude fragments of ST receptor binding peptides with deletions and/orinsertions and/or conservative substitutions.

In some embodiments, ST receptor binding peptides comprise D aminoacids. As used herein, the term “D amino acid peptides” is meant torefer to ST receptor binding peptides, fragments or derivatives whichcomprise at least one and preferably a plurality of D amino acids whichare capable of binding to the ST receptor. The use of D amino acidpeptides is desirable as they are less vulnerable to degradation andtherefore have a longer half-life. D amino acid peptides comprisingmostly all or consisting of D amino acids may comprise amino acidsequences in the reverse order of ST receptor binding peptides which aremade up of L amino acids.

In some embodiments, ST receptor binding peptides, including D aminoacid peptides, are conformationally restricted to present and maintainthe proper structural conformation for binding to the ST receptor. Thecompositions may comprise additional amino acid residues required toachieve proper three dimensional conformation including residues whichfacilitate circularization or desired folding.

It is preferred that the ST receptor ligand used as the ST receptorbinding moiety be as small as possible. Thus it is preferred that the STreceptor ligand be a non-peptide small molecule or small peptide,preferably less than 25 amino acids, more preferably less than 20 aminoacids. In some embodiments, the ST receptor ligand which constitute theST receptor binding moiety of a conjugated composition is less than 15amino acids. ST receptor binding peptide comprising less than 10 aminoacids and ST receptor binding peptide less than 5 amino acids may beused as ST binding moieties according to the present invention. It iswithin the scope of the present invention to include larger moleculeswhich serve as ST receptor binding moieties including, but not limitedto molecules such as antibodies, FAbs and F(Ab)2s which specificallybind to ST receptor.

An assay may be used to test both peptide and non-peptide compositionsto determine whether or not they are ST receptor ligands or, to testconjugated compositions to determine if they possess ST receptor bindingactivity. Such compositions that specifically bind to ST receptors canbe identified by a competitive binding assay. The competitive bindingassay is a standard technique in pharmacology which can be readilyperformed by those having ordinary skill in the art using readilyavailable starting materials. Competitive binding assays have been shownto be effective for identifying compositions that specifically bind toST receptors. Briefly, the assay consists of incubating a preparation ofST receptors (e.g. intestinal membranes from rat intestine, humanintestine, T84 cells) with a constant concentration (1×10⁻¹⁰ M to5×10⁻¹⁰ M) of ¹²⁵I-ST (any ST receptor ligand such as native STs SEQ IDNO:2, SEQ ID NO:3 or SEQ ID NO:5 may be used) and a known concentrationof a test compound. As a control, a duplicate preparation of STreceptors are incubated with a duplicate concentration of ¹²⁵I-ST in theabsence of test compound. Assays are incubated to equilibrium (2 hours)and the amount of ¹²⁵I-ST bound to receptors is quantified by standardtechniques. The ability of the test compound to bind to receptors ismeasured as its ability to prevent (compete with) the ¹²⁵I-ST frombinding. Thus, in assays containing the test compound which bind to thereceptor, there will be less radioactivity associated with thereceptors. This assay, which is appropriate for determining the abilityof any molecule to bind to ST receptors, is a standard competitivebinding assay which can be readily employed by those having ordinaryskill in the art using readily available starting materials.

ST may be isolated from natural sources using standard techniques.Additionally, ST receptor binding peptides and conjugated compositionsor portions thereof which are peptides may be prepared routinely by anyof the following known techniques.

ST receptor binding peptides and conjugated compositions or portionsthereof which are peptides may be prepared using the solid-phasesynthetic technique initially described by Merrifield, in J. Am. Chem.Soc., 15:2149-2154 (1963). Other peptide synthesis techniques may befound, for example, in M. Bodanszky et al., (1976) Peptide Synthesis,John Wiley & Sons, 2d Ed.; Kent and Clark-Lewis in Synthetic Peptides inBiology and Medicine, p. 295-358, eds. Alitalo, K., et al. SciencePublishers, (Amsterdam, 1985); as well as other reference works known tothose skilled in the art. A summary of peptide synthesis techniques maybe found in J. Stuart and J. D. Young, Solid Phase Peptide Synthelia,Pierce Chemical Company, Rockford, Ill. (1984), which is incorporatedherein by reference. The synthesis of peptides by solution methods mayalso be used, as described in The Proteins, Vol. II, 3d Ed., p. 105-237,Neurath, H. et al., Eds., Academic Press, New York, N.Y. (1976).Appropriate protective groups for use in such syntheses will be found inthe above texts, as well as in J. F. W. McOmie, Protective Groups inOrganic Chemistry, Plenum Press, New York, N.Y. (1973), which isincorporated herein by reference. In general, these synthetic methodsinvolve the sequential addition of one or more amino acid residues orsuitable protected amino acid residues to a growing peptide chain.Normally, either the amino or carboxyl group of the first amino acidresidue is protected by a suitable, selectively removable protectinggroup. A different, selectively removable protecting group is utilizedfor amino acids containing a reactive side group, such as lysine.

Using a solid phase synthesis as an example, the protected orderivatized amino acid is attached to an inert solid support through itsunprotected carboxyl or amino group. The protecting group of the aminoor carboxyl group is then selectively removed and the next amino acid inthe sequence having the complementary (amino or carboxyl) group suitablyprotected is admixed and reacted with the residue already attached tothe solid support. The protecting group of the amino or carboxyl groupis then removed from this newly added amino acid residue, and the nextamino acid (suitably protected) is then added, and so forth. After allthe desired amino acids have been linked in the proper sequence, anyremaining terminal and side group protecting groups (and solid support)are removed sequentially or concurrently, to provide the final peptide.The peptide of the invention are preferably devoid of benzylated ormethylbenzylated amino acids. Such protecting group moieties may be usedin the course of synthesis, but they are removed before the peptides areused. Additional reactions may be necessary, as described elsewhere, toform intramolecular linkages to restrain conformation.

ST receptor binding peptides and conjugated compositions or portionsthereof which are peptides may also be prepared by recombinant DNAtechniques. Provision of a suitable DNA sequence encoding the desiredpeptide permits the production of the peptide using recombinanttechniques now known in the art. The coding sequence can be obtainedfrom natural sources or synthesized or otherwise constructed usingwidely available starting materials by routine methods. When the codingDNA is prepared synthetically, advantage can be taken of known codonpreferences of the intended host where the DNA is to be expressed.

To produce an ST receptor binding peptide which occurs in nature, onehaving ordinary skill in the art can, using well-known techniques,obtain a DNA molecule encoding the ST receptor binding peptides from thegenome of the organism that produces the ST receptor binding peptide andinsert that DNA molecule into a commercially available expression vectorfor use in well-known expression systems.

Likewise, one having ordinary skill in the art can, using well knowntechniques, combine a DNA molecule that encodes an ST receptor bindingpeptide, such as SEQ ID NO:1 and SEQ ID NO:4, which can be obtained fromthe genome of the organism that produces the ST, with DNA that encodes atoxin, another active agent that is a peptide or additionally, any otheramino acid sequences desired to be adjacent to the ST receptor bindingpeptide amino acid sequence in a single peptide and insert that DNAmolecule into a commercially available expression vector for use inwell-known expression systems.

For example, the commercially available plasmid pSE420 (Invitrogen, SanDiego, Calif.) may be used for recombinant production in E. coli. Thecommercially available plasmid pYES2 (Invitrogen, San Diego, Calif.) maybe used for production in S. cerevisiae strains of yeast. Thecommercially available MaxBac™ (Invitrogen, San Diego, Calif.) completebaculovirus expression system may be used for production in insectcells. The commercially available plasmid pcDNA I (Invitrogen, SanDiego, Calif.) may be used for production in mammalian cells such asChinese Hamster Ovary cells.

One having ordinary skill in the art may use these or other commerciallyavailable expression vectors and systems or produce vectors usingwell-known methods and readily available starting materials. Expressionsystems containing the requisite control sequences, such as promotersand polyadenylation signals, and preferably enhancers, are readilyavailable and known in the art for a variety of hosts. See e.g.,Sambrook et al., Molecular Cloning a Laboratory Manual, Second Ed. ColdSpring Harbor Press (1989). Thus, the desired proteins can be preparedin both prokaryotic and eukaryotic systems, resulting in a spectrum ofprocessed forms of the protein.

The most commonly used prokaryotic system remains E. coli, althoughother systems such as B. subtilis and Pseudomonas are also useful.Suitable control sequences for prokaryotic systems include bothconstitutive and inducible promoters including the lac promoter, the trppromoter, hybrid promoters such as tac promoter, the lambda phage Plpromoter. In general, foreign proteins may be produced in these hostseither as fusion or mature proteins. When the desired sequences areproduced as mature proteins, the sequence produced may be preceded by amethionine which is not necessarily efficiently removed. Accordingly,the peptides and proteins claimed herein may be preceded by anN-terminal Met when produced in bacteria. Moreover, constructs may bemade wherein the coding sequence for the peptide is preceded by anoperable signal peptide which results in the secretion of the protein.When produced in prokaryotic hosts in this matter, the signal sequenceis removed upon secretion.

A wide variety of eukaryotic hosts are also now available for productionof recombinant foreign proteins. As in bacteria, eukaryotic hosts may betransformed with expression systems which produce the desired proteindirectly, but more commonly signal sequences are provided to effect thesecretion of the protein. Eukaryotic systems have the additionaladvantage that they are able to process introns which may occur in thegenomic sequences encoding proteins of higher organisms. Eukaryoticsystems also provide a variety of processing mechanisms which result in,for example, glycosylation, carboxy-terminal amidation, oxidation orderivatization of certain amino acid residues, conformational control,and so forth.

Commonly used eukaryotic systems include, but are not limited to, yeast,fungal cells, insect cells, mammalian cells, avian cells, and cells ofhigher plants. Suitable promoters are available which are compatible andoperable for use in each of these host types as well as are terminationsequences and enhancers, as e.g. the baculovirus polyhedron promoter. Asabove, promoters can be either constitutive or inducible. For example,in mammalian systems, the mouse metallothionene promoter can be inducedby the addition of heavy metal ions.

The particulars for the construction of expression systems suitable fordesired hosts are known to those in the art. For recombinant productionof the protein, the DNA encoding it is suitably ligated into theexpression vector of choice and then used to transform the compatiblehost which is then cultured and maintained under conditions whereinexpression of the foreign gene takes place. The protein of the presentinvention thus produced is recovered from the culture, either by lysingthe cells or from the culture medium as appropriate and known to thosein the art.

One having ordinary skill in the art can, using well-known techniques,isolate the protein that is produced.

According to the present invention, the active moiety may be atherapeutic agent or an imaging agent. One having ordinary skill in theart can readily recognize the advantages of being able to specificallytarget metastasized colorectal cells with an ST receptor ligand andconjugate such a ligand with many different active agents.

Chemotherapeutics useful as active moieties which when conjugated to anST receptor binding moiety are specifically delivered to metastasizedcolorectal cells are typically, small chemical entities produced bychemical synthesis. Chemotherapeutics include cytotoxic and cytostaticdrugs. Chemotherapeutics may include those which have other effects oncells such as reversal of the transformed state to a differentiatedstate or those which inhibit cell replication. Examples ofchemotherapeutics include common cytotoxic or cytostatic drugs such asfor example: methotrexate (amethopterin), doxorubicin (adrimycin),daunorubicin, cytosinarabinoside, etoposide, 5-4 fluorouracil,melphalan, chlorambucil, and other nitrogen mustards (e.g.cyclophosphamide), cis-platin vindesine (and other vinca alkaloids),mitomycin and bleomycin. Other chemotherapeutics include: purothionin(barley flour oligopeptide), macromomycin. 1,4-benzoquinone derivativesand trenimon.

Toxins are useful as active moieties. When a toxin is conjugated to anST receptor binding moiety, the conjugated composition is specificallydelivered to a metastasized colorectal cell by way of the ST receptorbinding moiety and the toxin moiety-kills the cell. Toxins are generallycomplex toxic products of various organisms including bacteria, plants,etc. Examples of toxins include but are not limited to: ricin, ricin Achain (ricin toxin), Pseudomonas exotoxin (PE) diphtheria toxin (DT),Clostridium perfringens phospholipase C (PLC), bovine pancreaticribonuclease (BPR), pokeweed antiviral protein (PAP), abrin, abrin Achain (abrin toxin), cobra venom factor (CVF), gelonin (GEL), saporin(SAP), modeccin, viscumin and volkensin. As discussed above, whenprotein toxins are employed with ST receptor binding peptides,conjugated compositions may be produced using recombinant DNAtechniques. Briefly, a recombinant DNA molecule can be constructed whichencodes both the ST receptor ligand and the toxin on a chimeric gene.When the chimeric gene is expressed, a fusion protein is produced whichincludes an ST receptor binding moiety and an active moiety. Proteintoxins are also useful to form conjugated compounds with ST receptorbinding peptides through non-peptidyl bonds.

In addition, there are other approaches for utilizing active agents forthe treatment of cancer. For example, conjugated compositions may beproduced which include an ST binding moiety and an active moiety whichis an active enzyme. The ST binding moiety specifically localizes theconjugated composition to the tumor cells. An inactive prodrug which canbe converted by the enzyme into an active drug is administered to thepatient. The prodrug is only converted to an active drug by the enzymewhich is localized to the tumor. An example of an enzyme/prodrug pairincludes alkaline phosphatase/etoposidephosphate. In such a case, thealkaline phosphatase is conjugated to an ST receptor binding ligand. Theconjugated compound is administered and localizes at the metastasizedcell. Upon contact with etoposidephosphate (the prodrug), theetoposidephosphate is converted to etoposide, a chemotherapeutic drugwhich is taken up by the cancer cell.

Radiosensitizing agents are substances that increase the sensitivity ofcells to radiation. Examples of radiosensitizing agents includenitroimidazoles, metronidazole and misonidazole (see: DeVita, V. T. Jr.in Harrison's Principles of Internal Medicine, p. 68, McGraw-Hill BookCo., N. Y. 1983, which is incorporated herein by reference). Theconjugated compound that comprises a radiosensitizing agent as theactive moiety is administered and localizes at the metastasized cell.Upon exposure of the individual to radiation, the radiosensitizing agentis “excited” and causes the death of the cell.

Radionuclides may be used in pharmaceutical compositions that are usefulfor radiotherapy or imaging procedures.

Examples of radionuclides useful as toxins in radiation therapy include:⁴⁷Sc, ⁶⁷Cu, ⁹⁰Y, ¹⁰⁹Pd, ¹²³I, ¹²⁵I, ¹³¹I, ¹⁸⁶Re, ¹⁸⁸Re, ¹⁹⁹Au, ²¹²At,²¹²Pb and ²¹²B. Other radionuclides which have been used by those havingordinary skill in the art include: ³²P and ³³P, ⁷¹Ge, ⁷⁷As, ¹⁰³Pb,¹⁰⁵Rh, ¹¹¹Ag, ¹¹⁹Sb, ¹²¹Sn, ¹³¹Cs, ¹⁴³Pr, ¹⁶¹Tb, ¹⁷⁷Lu, ¹⁹¹Os,^(193M)Pt, ¹⁹⁷Hg, all beta negative and/or auger emitters. Somepreferred radionuclides include: ⁹⁰Y, 131I, ²¹¹At and ²¹²Pb/²¹²Bi.

According to the present invention, the active moieties may be animaging agent. Imaging agents are useful diagnostic procedures as wellas the procedures used to identify the location of metastasized cells.Imaging can be performed by many procedures well-known to those havingordinary skill in the art and the appropriate imaging agent useful insuch procedures may be conjugated to an ST receptor ligand by well-knownmeans. Imaging can be performed, for example, by radioscintigraphy,nuclear magnetic resonance imaging (MRI) or computed tomography (CTscan). The most commonly employed radionuclide imaging agents includeradioactive iodine and indium. Imaging by CT scan may employ a heavymetal such as iron chelates. MRI scanning may employ chelates ofgadolinium or manganese. Additionally, positron emission tomography(PET) may be possible using positron emitters of oxygen, nitrogen, iron,carbon, or gallium. Example of radionuclides useful in imagingprocedures include: ⁴³K, ⁵²Fe, ⁵⁷Co, ⁶⁷Cu, ⁶⁷Ga, ⁶⁸Ga, ⁷⁷Br,⁸¹Rb/^(81M)Kr, ^(87M)Sr, ^(99M)Tc, ¹¹¹In, ^(113M)In, ¹²³I, ¹²⁵I, ¹²⁷Cs,¹²⁹Cs, ¹³¹I, ¹³²I, ¹⁹⁷Hg, ²⁰³Pb and ²⁰⁶Bi.

It is preferred that the conjugated compositions be non-immunogenic orimmunogenic at a very low level. Accordingly, it is preferred that theST receptor binding moiety be a small, poorly immunogenic ornon-immunogenic peptide or a non-peptide. Likewise, it is preferred thatthe active moiety be a small, poorly-immunogenic or non-immunogenicpeptide or a non-peptide. Native ST, being a small peptide, has beenshown to poorly immunogenic. (See: Klipstein, F. A. et al. (1982)Infect. Immun. 37:550-557; Giannella, R. A. et al. (1981) Infect. Immun.33:186; Burgess, M. N. et al. (1978) Infect. Immun. 21:60; Evans, D. G.et al. (1973) Infect. Immun. 7:873; Gyles, C. L. (1979) Ann. N.Y. Acad.Sci. 16:314; and Sack, R. B. (1975) Ann. Rev. Microbiol. 29:333.)Similarly, fragments and amino acid substitute derivatives of native STare poorly immunogenic. Accordingly, conjugated compositions whichinclude all or part of the native ST as an ST receptor binding moietyare generally poorly immunogenic to the extent that the native ST ispoorly immunogenic.

ST receptor ligands are conjugated to active agents by a variety ofwell-known techniques readily performed without undue experimentation bythose having ordinary skill in the art. The technique used to conjugatethe ST receptor ligand to the active agent is dependent upon themolecular nature of the ST receptor ligand and the active agent. Afterthe ST receptor ligand and the active agent are conjugated to form asingle molecule, assays may be performed to ensure that the conjugatedmolecule retains the activities of the moieties. The ST receptor bindingassay described above may be performed using the conjugated compound totest whether it is capable of binding to the ST receptor. Similarly, theactivity of the active moiety may be tested using various assays foreach respective type of active agent. Radionuclides retain thereactivity, i.e. their radioactivity, irrespective of conjugation. Withrespect to active agents which are toxins, drugs and targeting agents,standard assays to demonstrate the activity of unconjugated forms ofthese compounds may be used to confirm that the activity has beenretained.

Conjugation may be accomplished directly between the ST receptor ligandand the active agent or linking, intermediate molecular groups may beprovided between the ST receptor ligand and the active agent.Crosslinkers are particularly useful to facilitate conjugation byproviding attachment sites for each moiety. Crosslinkers may includeadditional molecular groups which serve as spacers to separate themoieties from each other to prevent either from interfering with theactivity of the other.

In some preferred embodiments, the ST receptor ligand peptide is SEQ IDNO:2, SEQ ID NO:3, SEQ ID NOS:5-54 or fragments or derivatives thereof.It has been observed that conjugation to these molecules is preferablyperformed at the amino terminus of each respective peptide. In STreceptor ligand peptides comprising D amino acid sequences in theopposite order as SEQ ID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54, conjugationpreferably is performed at the carboxy terminus.

One having ordinary skill in the art may conjugate an ST receptor ligandpeptide to a chemotherapeutic drug using well-known techniques. Forexample, Magerstadt, M. Antibody Conjugates and Malignant Disease.(1991) CRC Press, Boca Raton, USA, pp. 110-152) which is incorporatedherein by reference, teaches the conjugation of various cytostatic drugsto amino acids of antibodies. Such reactions may be applied to conjugatechemotherapeutic drugs to ST receptor ligands, including ST receptorbinding peptides, with an appropriate linker. ST receptor ligands whichhave a free amino group such as ST receptor binding peptides may beconjugated to active agents at that group. Most of the chemotherapeuticagents currently in use in treating cancer possess functional groupsthat are amenable to chemical crosslinking directly with proteins. Forexample, free amino groups are available on methotrexate, doxorubicin,daunorubicin, cytosinarabinoside, cis-platin, vindesine, mitomycin andbleomycin while free carboxylic acid groups are available onmethotrexate, melphalan, and chlorambucil. These functional groups, thatis free amino and carboxylic acids, are targets for a variety ofhomobifunctional and heterobifunctional chemical crosslinking agentswhich can crosslink these drugs directly to the single free amino groupof ST. For example, one procedure for crosslinking ST receptor ligandswhich have a free amino group such as ST receptor binding peptides, asfor example SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:5-54 to active agentswhich have a free amino group such as methotrexate, doxorubicin,daunorubicin, cytosinarabinoside, cis-platin, vindesine, mitomycin andbleomycin, or alkaline phosphatase, or protein- or peptide-based toxinemploys homobifunctional succinimidyl esters, preferably with carbonchain spacers such as disuccinimidyl suberate (Pierce Co, Rockford,Ill.). In the event that a cleavable conjugated compound is required,the same protocol would be employed utilizing 3,3′-dithiobis(sulfosuccinimidylpropionate; Pierce Co.).

In order to conjugate an ST receptor ligand peptide to a peptide-basedactive agent such as a toxin, the ST receptor ligand and the toxin maybe produced as a single, fusion protein either by standard peptidesynthesis or recombinant DNA technology, both of which can be routinelyperformed by those having ordinary skill in the art. Alternatively, twopeptides, the ST receptor ligand peptide and the peptide-based toxin maybe produced and/or isolated as separate peptides and conjugated usingcrosslinkers. As with conjugated compositions that containchemotherapeutic drugs, conjugation of ST receptor binding peptides andtoxins can exploit the ability to modify the single free amino group ofan ST receptor binding peptide while preserving the receptor-bindingfunction of this molecule.

One having ordinary skill in the art may conjugate an ST receptor ligandpeptide to a radionuclide using well-known techniques. For example,Magerstadt, M. (1991) Antibody Conjugates And Malignant Disease, CRCPress, Boca Raton, FLA,; and Barchel, S. W. and Rhodes, B. H., (1983)Radioimaging and Radiotherapy, Elsevier, NY, N.Y., each of which isincorporated herein by reference, teach the conjugation of varioustherapeutic and diagnostic radionuclides to amino acids of antibodies.Such reactions may be applied to conjugate radionuclides to ST receptorligand peptides or to ST receptor ligands including ST receptor ligandpeptides with an appropriate linker.

The present invention provides pharmaceutical compositions that comprisethe conjugated compounds of the invention and pharmaceuticallyacceptable carriers or diluents. The pharmaceutical composition of thepresent invention may be formulated by one having ordinary skill in theart. Suitable pharmaceutical carriers are described in Remington'sPharmaceutical Sciences, A. Osol, a standard reference text in thisfield, which is incorporated herein by reference. In carrying outmethods of the present invention, conjugated compounds of the presentinvention can be used alone or in combination with other diagnostic,therapeutic or additional agents. Such additional agents includeexcipients such as coloring, stabilizing agents, osmotic agents andantibacterial agents.

The conjugated compositions of the invention can be, for example,formulated as a solution, suspension or emulsion in association with apharmaceutically acceptable parenteral vehicle. Examples of suchvehicles are water, saline, Ringer's solution, dextrose solution, and 5%human serum albumin. Liposomes may also be used. The vehicle may containadditives that maintain isotonicity (e.g., sodium chloride, mannitol)and chemical stability (e.g., buffers and preservatives) The formulationis sterilized by commonly used techniques and is pyrogen free. Forexample, a parenteral composition suitable for administration byinjection is prepared by dissolving 1.5% by weight of active ingredientin 0.9% sodium chloride solution.

The pharmaceutical compositions according to the present invention maybe administered as either a single dose or in multiple doses. Thepharmaceutical compositions of the present invention may be administeredeither as individual therapeutic agents or in combination with othertherapeutic agents. The treatments of the present invention may becombined with conventional therapies, which may be administeredsequentially or simultaneously.

The pharmaceutical compositions of the present invention may beadministered by any means that enables the conjugated composition toreach the targeted cells. In some embodiments, routes of administrationinclude those selected from the group consisting of intravenous,intraarterial, intraperitoneal, local administration into the bloodsupply of the organ in which the tumor resides or directly into thetumor itself. Intravenous administration is the preferred mode ofadministration. It may be accomplished with the aid of an infusion pump.

The dosage administered varies depending upon factors such as: thenature of the active moiety; the nature of the conjugated composition;pharmacodynamic characteristics; its mode and route of administration;age, health, and weight of the recipient; nature and extent of symptoms;kind of concurrent treatment; and frequency of treatment.

Because conjugated compounds are specifically targeted to cells with STreceptors, conjugated compounds which comprise chemotherapeutics ortoxins are administered in doses less than those which are used when thechemotherapeutics or toxins are administered as unconjugated activeagents, preferably in doses that contain up to 100 times less activeagent. In some embodiments, conjugated compounds which comprisechemotherapeutics or toxins are administered in doses that contain10-100 times less active agent as an active moiety than the dosage ofchemotherapeutics or toxins administered as unconjugated active agents.To determine the appropriate dose, the amount of compound is preferablymeasured in moles instead of by weight. In that way, the variable weightof different ST binding moieties does not affect the calculation.Presuming a one to one ratio of ST binding moiety to active moiety inconjugated compositions of the invention, less moles of conjugatedcompounds may be administered as compared to the moles of unconjugatedcompounds administered, preferably up to 100 times less moles.

Typically, chemotherapeutic conjugates are administered intravenously inmultiple divided doses.

Up to 20 gm IV/dose of methotrexate is typically administered in anunconjugated form. When methotrexate is administered as the activemoiety in a conjugated compound of the invention, there is a 10-to100-fold dose reduction. Thus, presuming each conjugated compoundincludes one molecule of methotrexate conjugated to one ST receptorbinding moiety, of the total amount of conjugated compound administered,up to about 0.2-2.0 g of methotrexate is present and thereforeadministered. In some embodiments, of the total amount of conjugatedcompound administered, up to about 200 mg-2 g of methotrexate is presentand therefore administered.

Methotrexate has a molecular weight of 455. One mole of the STpeptide-methotrexate conjugate weighs between about 1755-2955 dependingon the ST peptide used. The effective dose range for STpeptide-methotrexate conjugate is between about 10 to 1000 mg. In someembodiments, dosages of 50 to 500 mg of ST peptide-methotrexateconjugate are administered. In some embodiments, dosages of 80 to 240 mgof ST peptide-methotrexate conjugate are administered.

Doxorubicin and daunorubicin each weigh about 535. Thus, STpeptide-doxorubicin conjugates and ST peptide-daunorubicin conjugateseach have molecular weights of between about 1835-2553.5. Presuming eachconjugated compound includes one molecule of doxorubicin or daunorubicinconjugated to one ST receptor binding moiety, the effective dose rangefor ST peptide-doxorubicin conjugate or ST peptide-daunorubicinconjugate is between about 40 to 4000 mg. In some embodiments, dosagesof 100 to 1000 mg of ST peptide-doxorubicin conjugate or STpeptide-daunorubicin conjugate are administered. In some embodiments,dosages of 200 to 600 mg of ST peptide-doxorubicin conjugate or STpeptide-daunorubicin conjugate are administered.

Toxin-containing conjugated compounds are formulated for intravenousadministration. Using this approach, up to 6 nanomoles/kg of body weightof toxin have been administered as a single dose with marked therapeuticeffects in patients with melanoma (Spitler L. E., et al. (1987) CancerRes. 47:1717). In some embodiments, up to about 11 micrograms of STpeptide-toxin conjugated compound/kg of body weight may be administeredfor therapy.

Presuming each conjugated compound includes one molecule of ricin toxinA chain conjugated to an ST receptor binding moiety, conjugatedcompositions comprising ricin toxin A chain are administered in doses inwhich the proportion by weight of ricin toxin A chain is 1-500 μg of thetotal weight of the conjugated compound administered. In some preferredembodiments, conjugated compositions comprising ricin toxin A chain areadministered in doses in which the proportion by weight of ricin toxin Achain is 10-100 μg of the total weight of the conjugated compoundadministered. In some preferred embodiments, conjugated compositionscomprising ricin toxin A chain are administered in doses in which theproportion by weight of ricin toxin A chain is 2-50 μg of the totalweight of the conjugated compound administered. The molecular weight ofricin toxin A chain is 32,000. Thus, a conjugated compound that containsricin A chain linked to an ST peptide has a molecular weight of about33,300-34,500. The range of doses of such conjugated compounds to beadministered are 1 to 500 μg. In some embodiments, 10 to 100 μg of suchconjugated compounds are administered. In some embodiments, 20 to 50 μgof such conjugated compounds are administered.

Presuming each conjugated compound includes one molecule of diphtheriatoxin A chain conjugated to an ST receptor binding moiety, conjugatedcompositions comprising diphtheria toxin A chain are administered indoses in which the proportion by weight of diphtheria toxin A chain is1-500 μg of the total weight of the conjugated compound administered. Insome preferred embodiments, conjugated compositions comprisingdiphtheria toxin A chain are administered in doses in which theproportion by weight of diphtheria toxin A chain is 10-100 μg of thetotal weight of the conjugated compound administered. In some preferredembodiments, conjugated compositions comprising diphtheria toxin A chainare administered in doses in which the proportion by weight ofdiphtheria toxin A chain is 40-80 μg of the total weight of theconjugated compound administered. The molecular weight of diphtheriatoxin A chain is 66,600. Thus, a conjugated compound that containsdiphtheria A chain linked to an ST peptide has a molecular weight ofabout 67,900-69,100. The range of doses of such conjugated compounds tobe administered tested are 1 to 500 μg. In some embodiments, 10 to 100μg of such conjugated compounds are administered. In some embodiments,40 to 80 μg of such conjugated compounds are administered.

Presuming each conjugated compound includes one molecule of Pseudomonasexotoxin conjugated to an ST receptor binding moiety, conjugatedcompositions comprising Pseudomonas exotoxin are administered in dosesin which the proportion by weight of Pseudomonas exotoxin is 0.01-100 μgof the total weight of the conjugated compound administered. In somepreferred embodiments, conjugated compositions comprising Pseudomonasexotoxin are administered in doses in which the proportion by weight ofPseudomonas exotoxin is 0.1-10 μg of the total weight of the conjugatedcompound administered. In some preferred embodiments, conjugatedcompositions comprising Pseudomonas exotoxin are administered in dosesin which the proportion by weight of Pseudomonas exotoxin is 0.3-2.2 μgof the total weight of the conjugated compound administered. Themolecular weight of Pseudomonas exotoxin is 22,000. Thus, a conjugatedcompound that contains Pseudomonas exotoxin linked to an ST peptide hasa molecular weight of about 23,300-24,500. The range of doses of suchconjugated compounds to be administered tested are 0.01 to 100 μg. Insome embodiments, 0.1 to 10 μg of such conjugated compounds areadministered. In some embodiments, 0.3 to 2.2 μg of such conjugatedcompounds are administered.

To dose conjugated compositions comprising ST receptor binding moietieslinked to active moieties that are radioisotopes in pharmaceuticalcompositions useful as imaging agents, it is presumed that each STreceptor binding moiety is linked to one radioactive active moiety. Theamount of radioisotope to be administered is dependent upon theradioisotope. Those having ordinary skill in the art can readilyformulate the amount of conjugated compound to be administered basedupon the specific activity and energy of a given radionuclide used as anactive moiety. Typically 0.1-100 millicuries per dose of imaging agent,preferably 1-10 millicuries, most often 2-5 millicuries areadministered. Thus, pharmaceutical compositions according to the presentinvention useful as imaging agents which comprise conjugatedcompositions comprising an ST receptor binding moiety and a radioactivemoiety comprise 0.1-100 millicuries, in some embodiments preferably 1-10millicuries, in some embodiments preferably 2-5 millicuries, in someembodiments more preferably 1-5 millicuries. Examples of dosagesinclude: ¹³¹I=between about 0.1-100 millicuries per dose, in someembodiments preferably 1-10 millicuries, in some embodiments 2-5millicuries, and in some embodiments about 4 millicuries; ¹¹¹In=betweenabout 0.1-100 millicuries per dose, in some embodiments preferably 1-10millicuries, in some embodiments 1-5 millicuries, and in someembodiments about 2 millicuries; ^(99m)Tc=between about 0.1-100millicuries per dose, in some embodiments preferably 5-75 millicuries,in some embodiments 10-50 millicuries, and in some embodiments about 27millicuries. Depending upon the specific activity of the radioactivemoiety and the weight of the ST receptor binding moiety the dosagedefined by weight varies. ST peptides have molecular weights of betweenabout 1300-2500. In the pharmaceutical composition comprising an STpeptide linked to a single ¹³¹I in which the specific activity of¹³¹I-ST peptide is about 2000 Ci/mmol, administering the dose of 0.1-100millicuries is the equivalent of 0.1-100 μg ¹³¹I-ST peptide,administering the dose of 1-10 millicuries is the equivalent of 1-10 μgof ¹³¹I-ST peptide, administering the dose of 2-5 millicuries isequivalent to giving 2-5 μg of ¹³¹I-ST peptide and administering thedose of 1-5 millicuries is equivalent to giving 1-5 μg of ¹³¹I-STpeptide. In the pharmaceutical composition comprising an ST peptidelinked to a single ¹¹¹In in which the specific activity of ¹¹¹In-STpeptide is about 1 Ci/mmol, administering the dose of 0.1-100millicuries is the equivalent of 0.2-200 mg ¹¹¹In-ST peptide,administering the dose of 1-10 millicuries is the equivalent of 2-20 mgof ¹¹¹In-ST peptide, administering the dose of 2-5 millicuries isequivalent to giving 4-10 mg of ¹¹¹In-ST peptide and administering thedose of 1-5 millicuries is equivalent to giving 2-10 mg of ¹¹¹In-STpeptide.

To dose conjugated compositions comprising ST receptor binding moietieslinked to active moieties that are radioisotopes in pharmaceuticalcompositions useful as therapeutic agents, it is presumed that each STreceptor binding moiety is linked to one radioactive active moiety. Theamount of radioisotope to be administered is dependent upon theradioisotope. Those having ordinary skill in the art can readilyformulate the amount of conjugated compound to be administered basedupon the specific activity and energy of a given radionuclide used as anactive moiety. For therapeutics that comprise ¹³¹I, between 10-1000 nM,preferably 50-500, more preferably about 300 nanomoles of ¹³¹I at thetumor, per gram of tumor, is desirable. Thus, if there is about 1 gramof tumor, and about 0.1% of the administered dose binds to the tumor,0.5-100 mg of ¹³¹I-ST peptide conjugated compound is administered. Insome embodiments, 1 to 50 mg of ¹³¹I-ST peptide conjugated compound isadministered. In some embodiments, 5 to 10 mg of ¹³¹I-ST peptideconjugated compound is administered. Wessels B. W. and R. D. Rogus(1984) Med. Phys. 11:638 and Kwok, C. S. et al. (1985) Med. Phys.12:405, both of which are incorporated herein by reference, disclosedetailed dose calculations for diagnostic and therapeutic conjugateswhich may be used in the preparation of pharmaceutical compositions ofthe present invention which include radioactive conjugated compounds.

One aspect of the present invention relates to a method of treatingindividuals suspected of suffering from metastasized colorectal cancer.Such individuals may be treated by administering to the individual apharmaceutical composition that comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radiostable therapeutic agent. In some embodiments of the presentinvention, the pharmaceutical composition comprises a pharmaceuticallyacceptable carrier or diluent and a conjugated compound that comprisesan ST receptor binding moiety and an active moiety wherein the activemoiety is a radiostable active agent and the ST receptor binding moietyis a peptide. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radiostable active agent and the ST receptor binding moiety isselected from the group consisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ IDNOS:5-54 and fragments and derivatives thereof. In some embodiments ofthe present invention, the pharmaceutical composition comprises apharmaceutically acceptable carrier or diluent and a conjugated compoundthat comprises an ST receptor binding moiety and an active moietywherein the active moiety is a radiostable active agent and the STreceptor binding moiety is selected from the group consisting of: SEQ IDNO:2, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:54. In someembodiments of the present invention, the pharmaceutical compositioncomprises a pharmaceutically acceptable carrier or diluent and aconjugated compound that comprises an ST receptor binding moiety and anactive moiety wherein the active moiety is a radiostable therapeuticagent. In some embodiments of the present invention, the pharmaceuticalcomposition comprises a pharmaceutically acceptable carrier or diluentand a conjugated compound that comprises an ST receptor binding moietyand an active moiety wherein the active moiety is a radiostable activeagent selected from the group consisting of: methotrexate, doxorubicin,daunorubicin, cytosinarabinoside, etoposide, 5-4 fluorouracil,melphalan, chlorambucil, cis-platin, vindesine, mitomycin, bleomycin,purothionin, macromomycin, 1,4-benzoquinone derivatives, trenimon,ricin, ricin A chain, Pseudomonas exotoxin, diphtheria toxin,Clostridium perfringens phospholipase C, bovine pancreatic ribonuclease,pokeweed antiviral protein, abrin, abrin A chain, cobra venom factor,gelonin, saporin, modeccin, viscumin, volkensin, alkaline phosphatase,nitroimidazole, metronidazole and misonidazole. In some embodiments ofthe present invention, the pharmaceutical composition comprises apharmaceutically acceptable carrier or diluent and a conjugated compoundthat comprises an ST receptor binding moiety and an active moietywherein the ST receptor binding moiety is selected from the groupconsisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 and fragmentsand derivatives thereof and the active moiety is a radiostable activeagent selected from the group consisting of: methotrexate, doxorubicin,daunorubicin, cytosinarabinoside, etoposide, 5-4 fluorouracil,melphalan, chlorambucil, cis-platin, vindesine, mitomycin, bleomycin,purothionin, macromomycin, 1,4-benzoquinone derivatives, trenimon,ricin, ricin A chain, Pseudomonas exotoxin, diphtheria toxin,Clostridium perfringens phospholipase C, bovine pancreatic ribonuclease,pokeweed antiviral protein, abrin, abrin A chain, cobra venom factor,gelonin, saporin, modeccin, viscumin, volkensin, alkaline phosphatase,nitroimidazole, metronidazole and misonidazole. In some embodiments ofthe present invention, the pharmaceutical composition comprises apharmaceutically acceptable carrier or diluent and a conjugated compoundthat comprises an ST receptor binding moiety and an active moietywherein the active moiety is a radiostable active agent selected fromthe group consisting of: methotrexate, doxorubicin, daunorubicin,cytosinarabinoside, cis-platin, vindesine, mitomycin and bleomycin,alkaline phosphatase, ricin A chain, Pseudomonas exotoxin and diphtheriatoxin. In some embodiments of the present invention, the pharmaceuticalcomposition comprises a pharmaceutically acceptable carrier or diluentand a conjugated compound that comprises an ST receptor binding moietyand an active moiety wherein the ST receptor binding moiety is selectedfrom the group consisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:5, SEQID NO:6 and SEQ ID NO:54 and the active moiety is a radiostable activeagent selected from the group consisting of: methotrexate, doxorubicin,daunorubicin, cytosinarabinoside, cis-platin, vindesine, mitomycin andbleomycin, alkaline phosphatase, ricin A chain, Pseudomonas exotoxin anddiphtheria toxin. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a radiostable conjugated compound described inExample 1. The individual being treated may be diagnosed as havingmetastasized colorectal cancer or may be diagnosed as having localizedcolorectal cancer and may undergo the treatment proactively in the eventthat there is some metastasis as yet undetected. The pharmaceuticalcomposition contains a therapeutically effective amount of theconjugated composition. A therapeutically effective amount is an amountwhich is effective to cause a cytotoxic or cytostatic effect onmetastasized colorectal cancer cells without causing lethal side effectson the individual.

One aspect of the present invention relates to a method of treatingindividuals suspected of suffering from metastasized colorectal cancer.Such individuals may be treated by administering to the individual apharmaceutical composition that comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radioactive. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radioactive and the ST receptor binding moiety is a peptide. Insome embodiments of the present invention, the pharmaceuticalcomposition comprises a pharmaceutically acceptable carrier or diluentand a conjugated compound that comprises an ST receptor binding moietyand an active moiety wherein the active moiety is a radioactive and theST receptor binding moiety is selected from the group consisting of: SEQID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 and fragments and derivativesthereof. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radioactive and the ST receptor binding moiety is selected from thegroup consisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO: 6and SEQ ID NO:54. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radioactive agent selected from the group consisting of: ⁴⁷SC,⁶⁷Cu, ⁹⁰Y, ¹⁰⁹Pd, ¹²³I, ₁₂₅I, ¹³¹I, ¹⁸⁶Re, ¹⁸⁸Re, ¹⁹⁹Au, ²¹¹At, ²¹²Pb,²¹²B, ³²P and ³³P, ⁷¹Ge, ⁷⁷As, ¹⁰³Pb, ¹⁰⁵Rh, ¹¹¹Ag, ¹¹⁹Sb, ¹²¹Sn, ¹³¹Cs,143Pr, ¹⁶¹Tb, ¹⁷⁷Lu, ¹⁹¹Os, ^(193M)Pt and ¹⁹⁷Hg. In some embodiments ofthe present invention, the pharmaceutical composition comprises apharmaceutically acceptable carrier or diluent and a conjugated compoundthat comprises an ST receptor binding moiety and an active moietywherein the ST receptor binding moiety is selected from the groupconsisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 and fragmentsand derivatives thereof and the active moiety is a radioactive agentselected from the group consisting of: ⁴⁷Sc, ⁶⁷Cu, ⁹⁰Y, ¹⁰⁹Pd, ¹²³I,¹²⁵I, ¹³¹I, ¹⁸⁶Re, ¹⁸⁸Re, ¹⁹⁹Au, ²¹¹At, ²¹²Pb, ²¹²B, ³²P and ³³P, ⁷¹Ge,⁷⁷As, ¹⁰³Pb, ¹⁰⁵Rh, ¹¹¹Ag, ¹¹⁹Sb, ¹²¹Sn, ¹³¹Cs, ¹⁴³Pr, ¹⁶¹Tb, ¹⁷⁷Lu,¹⁹¹Os, ^(193M)Pt, ¹⁹⁷Hg, ³²P and ³³P, ⁷¹Ge, ⁷⁷As, ¹⁰³Pb, ¹⁰⁵Rh, ¹¹¹Ag,¹¹⁹Sb, ¹²¹Sn, ¹³¹Cs, ¹⁴³Pr, ¹⁶¹Tb, ¹⁷⁷Lu, ¹⁹⁷Hg, all beta negativeand/or auger emitters. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radioactive agent selected from the group consisting of: ⁴⁷Sc,⁶⁷Cu, ⁹⁰Y, ¹⁰⁹Pd, ¹²³I, ¹²⁵I, ¹³¹I, ¹⁸⁶Re, ¹⁸⁸Re, ¹⁹⁹Au, ²¹¹At, ²¹²Pb,²¹²B ³²P and ³³P, ⁷¹Ge, ⁷⁷As, ¹⁰³Pb, ¹⁰⁵Rh, ¹¹¹Ag, ¹¹⁹Sb, ¹²¹Sn, ¹³¹Cs,¹⁴³Pr, ¹⁶¹Tb, ¹⁷⁷Lu, ¹⁹¹Os, ^(193M)Pt and ¹⁹⁷Hg. In some embodiments ofthe present invention, the pharmaceutical composition comprises apharmaceutically acceptable carrier or diluent and a conjugated compoundthat comprises an ST receptor binding moiety and an active moietywherein the ST receptor binding moiety is selected from the groupconsisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:6 andSEQ ID NO:54 and the active moiety is a radioactive agent selected fromthe group consisting of: ⁴⁷Sc, ⁶⁷Cu, ⁹⁰Y, ¹⁰⁹Pd, ¹²³I, ¹²⁵I, ¹³¹I,¹⁸⁶Re, ¹⁸⁸Re, ¹⁹⁹Au, ²¹¹At, ²¹²Pb, ²¹²B, ³²P and ³³P, ⁷¹Ge, ⁷⁷As, ¹⁰³Pb,¹⁰⁵Rh, ¹¹¹ Ag, ¹¹⁹Sb, ¹²¹Sn, ¹³¹Cs, ¹⁴³Pr, ¹⁶¹Tb, ¹⁷⁷Lu, ¹⁹¹Os,^(193M)Pt and ¹⁹⁷Hg. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a radioactive conjugated compound described inExample 1. The individual being treated may be diagnosed as havingmetastasized colorectal cancer or may be diagnosed as having localizedcolorectal cancer and may undergo the treatment proactively in the eventthat there is some metastasis as yet undetected. The pharmaceuticalcomposition contains a therapeutically effective amount of theconjugated composition. A therapeutically effective amount is an amountwhich is effective to cause a cytotoxic or cytostatic effect onmetastasized colorectal cancer cells without causing lethal side effectson the individual.

One aspect of the present invention relates to a method of detectingmetastasized colorectal cancer cells in an individual suspected ofsuffering from metastasized colorectal cancer by radioimaging. Suchindividuals may be diagnosed as suffering from metastasized colorectalcancer and the metastasized colorectal cancer cells may be detected byadministering to the individual, preferably by intravenousadministration, a pharmaceutical composition that comprises apharmaceutically acceptable carrier or diluent and a conjugated compoundthat comprises an ST receptor binding moiety and an active moietywherein the active moiety is a radioactive and detecting the presence ofa localized accumulation or aggregation of radioactivity, indicating thepresence of cells with ST receptors. In some embodiments of the presentinvention, the pharmaceutical composition comprises a pharmaceuticallyacceptable carrier or diluent and a conjugated compound that comprisesan ST receptor binding moiety and an active moiety wherein the activemoiety is a radioactive and the ST receptor binding moiety is a peptide.In some embodiments of the present invention, the pharmaceuticalcomposition comprises a pharmaceutically acceptable carrier or diluentand a conjugated compound that comprises an ST receptor binding moietyand an active moiety wherein the active moiety is a radioactive and theST receptor binding moiety is selected from the group consisting of: SEQID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 and fragments and derivativesthereof. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radioactive and the ST receptor binding moiety is selected from thegroup consisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:6and SEQ ID NO:54. In some embodiments of the present invention, thepharmaceutical composition comprises a pharmaceutically acceptablecarrier or diluent and a conjugated compound that comprises an STreceptor binding moiety and an active moiety wherein the active moietyis a radioactive agent selected from the group consisting of:radioactive heavy metals such as iron chelates, radioactive chelates ofgadolinium or manganese, positron emitters of oxygen, nitrogen, iron,carbon, or gallium, ⁴³K, ⁵²Fe, ⁵⁷Co, ⁶⁷Cu, ⁶⁷Ga, ⁶⁸Ga, ⁷⁷Br,⁸¹Rb/^(81M)Kr, ^(87M)Sr, ^(99M)Tc, ¹¹¹In, ^(113M)In, ¹²³I, ¹²⁵I, ¹²⁷Cs,¹²⁹Cs, ¹³¹I, ¹³²I, ¹⁹⁷Hg, ²⁰³Pb and ²⁰⁶Bi. In some embodiments of thepresent invention, the pharmaceutical composition comprises apharmaceutically acceptable carrier or diluent and a conjugated compoundthat comprises an ST receptor binding moiety and an active moietywherein the ST receptor binding moiety is selected from the groupconsisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 and fragmentsand derivatives thereof and the active moiety is a radioactive agentselected from the group consisting of: radioactive heavy metals such asiron chelates, radioactive chelates of gadolinium or manganese, positronemitters of oxygen, nitrogen, iron, carbon, or gallium, ⁴³K, ⁵²Fe, ⁵⁷Co,⁶⁷Cu, ⁶⁷Ga, ⁶⁸Ga, ⁷⁷Br, ⁸¹Rb/^(81M)Kr, ^(87M)Sr, ^(99M)Tc, ¹¹¹In,^(113M)In, ¹²³I, ¹²⁵I, ¹²⁷Cs, ¹²⁹Cs, ¹³¹I, ¹³²I, ¹⁹⁷Hg, ²⁰³Pb and ²⁰⁶Bi.In some embodiments of the present invention, the pharmaceuticalcomposition comprises a pharmaceutically acceptable carrier or diluentand a conjugated compound that comprises an ST receptor binding moietyand an active moiety wherein the active moiety is a radioactive agentselected from the group consisting of: ⁴³K, ⁵²Fe, ⁵⁷Co, ⁶⁷Cu, ⁶⁷Ga,⁶⁸Ga, ⁷⁷Br, ⁸¹Rb/^(81M)Kr, ^(87M)Sr, ^(99M)Tc, ¹¹¹In, ^(113M)In, ¹²³I,¹²⁵I, ¹²⁷Cs, ¹²⁹Cs, ¹³¹I, ¹³²I, ¹⁹⁷Hg, ²⁰³Pb and ²⁰⁶Bi. In someembodiments of the present invention, the pharmaceutical compositioncomprises a pharmaceutically acceptable carrier or diluent and aconjugated compound that comprises an ST receptor binding moiety and anactive moiety wherein the ST receptor binding moiety is selected fromthe group consisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:5, SEQ IDNO:6 and SEQ ID NO:54 and the active moiety is a radioactive agentselected from the group consisting of: ⁴³K, ⁵²Fe, ⁵⁷Co, ⁶⁷Cu, ⁶⁷Ga,⁶⁸Ga, ⁷⁷Br, ⁸¹Rb/^(81M)Kr, ^(87M)Sr, ^(99M)Tc, ¹¹¹In, ^(113M)In, ¹²³I,¹²⁵I, ¹²⁷Cs, ¹²⁹Cs, ¹³¹I, ¹³²I, ¹⁹⁷Hg, ²⁰³Pb and ²⁰⁶Bi. In someembodiments of the present invention, the pharmaceutical compositioncomprises a pharmaceutically acceptable carrier or diluent and aradioactive conjugated compound described in Example 1. The individualbeing treated may be diagnosed as having metastasized colorectal canceror may be diagnosed as having localized colorectal cancer and mayundergo the treatment proactively in the event that there is somemetastasis as yet undetected. The pharmaceutical composition contains adiagnostically effective amount of the conjugated composition. Adiagnostically effective amount is an amount which can be detected at asite in the body where cells with ST receptors are located withoutcausing lethal side effects on the individual.

Another aspect of the invention relates to unconjugated compositionswhich comprise an ST receptor binding ligand and an active agent. Forexample, liposomes are small vesicles composed of lipids. Drugs can beintroduced into the center of these vesicles. The outer shell of thesevesicles comprise an ST receptor binding ligand. Liposomes Volumes 1, 2and 3 CRC Press Inc. Boca Raton FLA, which is incorporated herein byreference, disclose preparation of liposome-encapsulated active agentswhich include targeting agents that correspond to ST receptor ligand inthe outer shell. Unconjugated compositions which comprise an ST receptorligand in the matrix of a liposome with an active agent inside includesuch compostions in which the ST receptor ligand is selected from thegroup consisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 andfragments and derivatives thereof and the active agent is selected fromthe group consisting of: methotrexate, doxorubicin, daunorubicin,cytosinarabinoside, etoposide, 5-4 fluorouracil, melphalan,chlorambucil, cis-platin, vindesine, mitomycin, bleomycin, purothionin,macromomycin, 1,4-benzoquinone derivatives, trenimon, ricin, ricin Achain, Pseudomonas exotoxin, diphtheria toxin, Clostridium perfringensphospholipase C, bovine pancreatic ribonuclease, pokeweed antiviralprotein, abrin, abrin A chain, cobra venom factor, gelonin, saporin,modeccin, viscumin, volkensin, alkaline phosphatase, nitroimidazole,metronidazole and misonidazole.

Another aspect of the invention relates to unconjugated and conjugatedcompositions which comprise an ST receptor ligand used to delivertherapeutic nucleic acid molecules to cells that comprise an ST receptorsuch as normal cells of the intestinal tract as well as metastasizedcolorectal cancer cells. In some embodiments, the genetic material isdelivered to metastasized tumor cells to produce an antigen that can betargeted by the immune system or to produce a protein which kills thecell or inhibits its proliferation. In some embodiments, the ST receptorligand is used to deliver nucleic acids that encode nucleic acidmolecules which replace defective endogenous genes or which encodetherapeutic proteins. In some embodiments, the ST receptor ligand isthus used to deliver the active agent specifically to the cells liningthe intestinal tract to treat diseases specific to this organ. Accordingto this aspect of the invention, compositions comprise nucleic acidmolecules which can replace defective genes. In some embodiments, thecompositions are used in gene therapy protocols to deliver toindividuals, genetic material needed and/or desired to make up for agenetic deficiency.

In some embodiments, the ST receptor ligand is combined with orincorporated into a delivery vehicle thereby converting the deliveryvehicle into a specifically targeted delivery vehicle. For example, anST receptor binding peptide may be integrated into the outer portion ofa viral particle making such a virus an ST receptor-bearing cellspecific virus. Similarly, the coat protein of a virus may be engineeredsuch that it is produced as a fusion protein which includes an active STreceptor binding peptide that is exposed or otherwise accessible on theoutside of the viral particle making such a virus an ST receptor-bearingcell-specific virus. In some embodiments, an ST receptor ligand may beintegrated or otherwise incorporated into the liposomes wherein the STreceptor ligand is exposed or otherwise accessible on the outside of theliposome making such liposomes specifically targeted to STreceptor-bearing cells.

The active agent in the conjugated or unconjugated compositionsaccording to this aspect of the invention is a nucleic acid molecule.The nucleic acid may be RNA or preferably DNA. In some embodiments, thenucleic acid molecule is an antisense molecule or encodes an antisensesequence whose presence in the cell inhibits production of anundesirable protein. In some embodiments, the nucleic acid moleculeencodes a ribozyme whose presence in the cell inhibits production of anundesirable protein. In some embodiments, the nucleic acid moleculeencodes a protein or peptide that is desirably produced in the cell. Insome embodiments, the nucleic acid molecule encodes a functional copy ofa gene that is defective in the targeted cell. The nucleic acid moleculeis preferably operably linked to regulatory elements needed to expressthe coding sequence in the cell.

Liposomes are small vesicles composed of lipids. Genetic constructswhich encode proteins that are desired to be expressed in STreceptor-bearing cells are introduced into the center of these vesicles.The outer shell of these vesicles comprise an ST receptor ligand, insome embodiments preferably an ST peptide. Liposomes Volumes 1, 2 and 3CRC Press Inc. Boca Raton FLA, which is incorporated herein byreference, disclose preparation of liposome-encapsulated active agentswhich include antibodies in the outer shell. In the present invention,an ST receptor ligand such as for example an ST peptide corresponds tothe antibodies in the outer shell. Unconjugated compositions whichcomprise an ST receptor ligand in the matrix of a liposome with anactive agent inside include such compositions in which the ST receptorligand is selected from the group consisting of: SEQ ID NO:2, SEQ IDNO:3, SEQ ID NOS:5-54 and fragments and derivatives thereof.

In one embodiment for example, cystic fibrosis, a genetic disease inwhich there is a mutation of a specific gene encoding a chloridetransport protein which ultimately produces abnormalities of function inmany systems, most notably in the respiratory and intestinal tract, istreated by gene therapy techniques using ST receptor ligands to deliverthe corrective gene to cells. Current therapy has been directed atreplacing the mutant gene in the respiratory system with the normal geneby targeting these genes directly to the cells lining the respiratorytract using viruses which bind only to those cells. Similarly, thenormal gene is packaged in liposomes targeted on their surface with STreceptor ligands and delivered to the intestinal tract. ST receptorligands specifically target and direct the liposomes containing thenormal gene to correct the lesion for cystic fibrosis to the specificcells lining the intestinal tract, from the duodenum to the rectum.Uptake of that genetic material by those cells should result in a cureof cystic fibrosis in the intestinal tract.

In another embodiment, the delivery of normal copies of the p53 tumorsuppressor gene to the intestinal tract is accomplished using STreceptor ligand to target the gene therapeutic. Mutations of the p53tumor suppressor gene appears to play a prominent role in thedevelopment of colorectal cancer in the intestinal tract. One approachto combatting this disease is the delivery of normal copies of this geneto the intestinal tract to cells expressing mutant forms of this gene.Genetic constructs that comprise normal p53 tumor suppressor genes areincorporated into liposomes that comprise an ST receptor ligand. Thecomposition is delivered to the intestinal tract. ST receptor bindingligands specifically target and direct the liposomes containing thenormal gene to correct the lesion created by mutation of p53 suppressorgene in intestinal cells.

Preparation of genetic constructs is with the skill of those havingordinary skill in the art. The present invention allows such constructto be specifically targeted by using the ST receptor ligands of thepresent invention. The compositions of the invention include an STreceptor ligand such as an ST peptide associated with a delivery vehicleand a gene construct which comprises a coding sequence for a proteinwhose production is desired in the cells of the intestinal tract linkedto necessary regulatory sequences for expression in the cells. Foruptake by cells of the intestinal tract, the compositions areadministered orally or by enema whereby they enter the intestinal tractand contact cells which comprise ST receptors. The delivery vehiclesassociate with the ST receptor by virtue of the ST receptor ligand andthe vehicle is internalized into the cell or the active agent/geneticconstruct is otherwise taken up by the cell. Once internalized, theconstruct can provide a therapeutic effect on the individual. One havingordinary skill in the art can readily formulate such compositions fororal or enema administration and determine the effective amount of suchcomposition to be administered to treat the disease or disorder.

The following examples are illustrative but are not meant to be limitingof the present invention.

EXAMPLES Example 1

The following are representative compounds according to the presentinvention. Whenever stated below, reference to a series of compounds isprovided for efficiency and is meant to name each compound in the seriesincluding all the compounds in numerical order, such as for example“3-D1 to 3-D16” is meant to refer to compounds 3-D1, 3-D2, 3-D3, 3-D4,3-D5, 3-D6, 3-D7, 3-D8, 3-D9, 3-D10, 3-D11, 3-D12, 3-D13, 3-D14, 3-D15and 3-D16. Likewise, whenever stated below, reference to a series of SEQID NO:'s is provided for efficiency and is meant to name each SEQ ID NO:in the series including the all SEQ ID NO:'s in numerical order, such asfor example SEQ ID NO:5 through SEQ ID NO:54 is meant to refer to SEQ IDNO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10,SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15,SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20,SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25,SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30,SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, SEQ ID NO:35,SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40,SEQ ID NO:41, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:45,SEQ ID NO:46, SEQ ID NO:47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:50,SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:53 and SEQ ID NO:54. Similarly,whenever stated below, reference to a series of compounds is providedfor efficiency and is meant to name each compound in the seriesincluding the all compounds in numerical order, such as for example“5-AP to 54-AP” is meant to refer to compounds 5-AP, 6-AP, 7-AP, 8-AP,9-AP, 10-AP, 11-AP, 12-AP, 13-AP, 14-AP, 15-AP, 16-AP, 17-AP, 18-AP,19-AP, 20-AP, 21-AP, 22-AP, 23-AP, 24-AP, 25-AP, 26-AP, 27-AP, 28-AP,29-AP, 30-AP, 31-AP, 32-AP, 33-AP, 34-AP, 35-AP, 36-AP, 37-AP, 38-AP,39-AP, 40-AP, 41-AP, 42-AP, 43-AP, 44-AP, 45-AP, 46-AP, 47-AP, 48-AP,49-AP, 50-AP, 51-AP, 52-AP, 53-AP and 54-AP.

Compound 2-D1 comprises methotrexate (amethopterin) conjugated to SEQ IDNO:2.

Compound 2-D2 comprises doxorubicin (adrimycin) conjugated to SEQ IDNO:2.

Compound 2-D3 comprises daunorubicin conjugated to SEQ ID NO:2.

Compound 2-D4 comprises cytosinarabinoside conjugated to SEQ ID NO:2.

Compound 2-D5 comprises etoposide conjugated to SEQ ID NO:2.

Compound 2-D6 comprises 5-4 fluorouracil conjugated to SEQ ID NO:2.

Compound 2-D7 comprises melphalan conjugated to SEQ ID NO:2.

Compound 2-D8 comprises chlorambucil conjugated to SEQ ID NO:2.

Compound 2-D9 comprises cyclophosphamide conjugated to SEQ ID NO:2.

Compound 2-D10 comprises cis-platin conjugated to SEQ ID NO:2.

Compound 2-D11 comprises vindesine conjugated to SEQ ID NO:2.

Compound 2-D12 comprises mitomycin conjugated to SEQ ID NO:2.

Compound 2-D13 comprises bleomycin conjugated to SEQ ID NO:2.

Compound 2-D14 comprises purothionin conjugated to SEQ ID NO:2.

Compound 2-D15 comprises macromomycin conjugated to SEQ ID NO:2.

Compound 2-D16 comprises trenimon conjugated to SEQ ID NO:2.

Compounds 3-D1 to 3-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 3-D1 to 3-D16 each comprise SEQ IDNO:3 as the ST receptor binding moiety.

Compounds 5-D1 to 5-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 5-D1 to 5-D16 each comprise SEQ IDNO:5 as the ST receptor binding moiety.

Compounds 6-D1 to 6-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 6-D1 to 6-D16 each comprise SEQ IDNO:6 as the ST receptor binding moiety.

Compounds 7-D1 to 7-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 7-D1 to 7-D16 each comprise SEQ IDNO:7 as the ST receptor binding moiety.

Compounds 8-D1 to 8-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 8-D1 to 8-D16 each comprise SEQ IDNO:8 as the ST receptor binding moiety.

Compounds 9-D1 to 9-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 9-D1 to 9-D16 each comprise SEQ IDNO:9 as the ST receptor binding moiety.

Compounds 10-D1 to 10-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 10-D1 to 10-D16 each comprise SEQ IDNO:2 as the ST receptor binding moiety.

Compounds 12-D1 to 12-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 12-D1 to 12-D16 each comprise SEQ IDNO:11 as the ST receptor binding moiety.

Compounds 12-D1 to 12-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 12-D1 to 12-D16 each comprise SEQ IDNO:12 as the ST receptor binding moiety.

Compounds 13-D1 to 13-D16 are the same as compounds 2-D1 to 2-D1,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 13-D1 to 13-D16 each comprise SEQ IDNO:13 as the ST receptor binding moiety.

Compounds 14-D1 to 14-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 14-D1 to 14-D16 each comprise SEQ IDNO:14 as the ST receptor binding moiety.

Compounds 15-D1 to 15-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 15-D1 to 15-D16 each comprise SEQ IDNO:15 as the ST receptor binding moiety.

Compounds 16-D1 to 16-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 16-D1 to 16-D16 each comprise SEQ IDNO:16 as the ST receptor binding moiety.

Compounds 17-D1 to 17-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 17-D1 to 17-D16 each comprise SEQ IDNO:17 as the ST receptor binding moiety.

Compounds 18-D1 to 18-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 18-D1 to 18-D16 each comprise SEQ IDNO:18 as the ST receptor binding moiety.

Compounds 19-D1 to 19-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 19-D1 to 19-D16 each comprise SEQ IDNO:19 as the ST receptor binding moiety.

Compounds 20-D1 to 20-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 20-D1 to 20-D16 each comprise SEQ IDNO:20 as the ST receptor binding moiety.

Compounds 22-D1 to 22-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 22-D1 to 22-D16 each comprise SEQ IDNO:21 as the ST receptor binding moiety.

Compounds 22-D1 to 22-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 22-D1 to 22-D16 each comprise SEQ IDNO:22 as the ST receptor binding moiety.

Compounds 23-D1 to 23-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 23-D1 to 23-D16 each comprise SEQ IDNO:23 as the ST receptor binding moiety.

Compounds 24-D1 to 24-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 24-D1 to 24-D16 each comprise SEQ IDNO:24 as the ST receptor binding moiety.

Compounds 25-D1 to 25-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 25-D1 to 25-D16 each comprise SEQ IDNO:25 as the ST receptor binding moiety.

Compounds 26-D1 to 26-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 26-D1 to 26-D16 each comprise SEQ IDNO:26 as the ST receptor binding moiety.

Compounds 27-D1 to 27-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 27-D1 to 27-D16 each comprise SEQ IDNO:27 as the ST receptor binding moiety.

Compounds 28-D1 to 28-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 28-D1 to 28-D16 each comprise SEQ IDNO:28 as the ST receptor binding moiety.

Compounds 29-D1 to 29-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 29-D1 to 29-D16 each comprise SEQ IDNO:29 as the ST receptor binding moiety.

Compounds 30-D1 to 30-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 30-D1 to 30-D16 each comprise SEQ IDNO:30 as the ST receptor binding moiety.

Compounds 32-D1 to 32-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 32-D1 to 32-D16 each comprise SEQ IDNO:31 as the ST receptor binding moiety.

Compounds 32-D1 to 32-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 32-D1 to 32-D16 each comprise SEQ IDNO:32 as the ST receptor binding moiety.

Compounds 33-D1 to 33-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 33-D1 to 33-D16 each comprise SEQ IDNO:33 as the ST receptor binding moiety.

Compounds 34-D1 to 34-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 34-D1 to 34-D16 each comprise SEQ IDNO:34 as the ST receptor binding moiety.

Compounds 35-D1 to 35-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 35-D1 to 35-D16 each comprise SEQ IDNO:35 as the ST receptor binding moiety.

Compounds 36-D1 to 36-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ 1S ID NO:2 as the STreceptor binding moiety, compounds 36-D1 to 36-D16 each comprise SEQ IDNO:36 as the ST receptor binding moiety.

Compounds 37-D1 to 37-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 37-D1 to 37-D16 each comprise SEQ IDNO:37 as the ST receptor binding moiety.

Compounds 38-D1 to 38-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 38-D1 to 38-D16 each comprise SEQ IDNO:38 as the ST receptor binding moiety.

Compounds 39-D1 to 39-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 39-D1 to 39-D16 each comprise SEQ IDNO:39 as the ST receptor binding moiety.

Compounds 40-D1 to 40-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 40-D1 to 40-D16 each comprise SEQ IDNO:40 as the ST receptor binding moiety.

Compounds 42-D1 to 42-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 42-D1 to 42-D16 each comprise SEQ IDNO:41 as the ST receptor binding moiety.

Compounds 42-D1 to 42-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 42-D1 to 42-D16 each comprise SEQ IDNO:42 as the ST receptor binding moiety.

Compounds 43-D1 to 43-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 43-D1 to 43-D16 each comprise SEQ IDNO:43 as the ST receptor binding moiety.

Compounds 44-D1 to 44-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 44-D1 to 44-D16 each comprise SEQ IDNO:44 as the ST receptor binding moiety.

Compounds 45-D1 to 45-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 45-D1 to 45-D16 each comprise SEQ IDNO:45 as the ST receptor binding moiety.

Compounds 46-D1 to 46-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 46-D1 to 46-D16 each comprise SEQ IDNO:46 as the ST receptor binding moiety.

Compounds 47-D1 to 47-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 47-D1 to 47-D16 each comprise SEQ IDNO:47 as the ST receptor binding moiety.

Compounds 48-D1 to 48-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 48-D1 to 48-D16 each comprise SEQ IDNO:48 as the ST receptor binding moiety.

Compounds 49-D1 to 49-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 49-D1 to 49-D16 each comprise SEQ IDNO:49 as the ST receptor binding moiety.

Compounds 50-D1 to 50-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 50-D1 to 50-D16 each comprise SEQ IDNO:50 as the ST receptor binding moiety.

Compounds 51-D1 to 51-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 51-D1 to 51-D16 each comprise SEQ IDNO:51 as the ST receptor binding moiety.

Compounds 52-D1 to 52-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 52-D1 to 52-D16 each comprise SEQ IDNO:52 as the ST receptor binding moiety.

Compounds 53-D1 to 53-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 53-D1 to 53-D16 each comprise SEQ IDNO:53 as the ST receptor binding moiety.

Compounds 54-D1 to 54-D16 are the same as compounds 2-D1 to 2-D16,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 54-D1 to 54-D1 G each comprise SEQ IDNO:54 as the ST receptor binding moiety.

Compound 2-T1 comprises ricin conjugated to SEQ ID NO:2.

Compound 2-T2 comprises ricin A chain (ricin toxin) conjugated to SEQ IDNO:2.

Compound 2-T3 comprises Pseudomonas exotoxin (PE) conjugated to SEQ IDNO:2.

Compound 2-T4 comprises diphtheria toxin (DT), conjugated to SEQ IDNO:2.

Compound 2-T5 comprises Clostridium perfringens phospholipase C (PLC)conjugated to SEQ ID NO:2.

Compound 2-T6 comprises bovine pancreatic ribonuclease (BPR) conjugatedto SEQ ID NO:2.

Compound 2-T7 comprises pokeweed antiviral protein (PAP) conjugated toSEQ ID NO:2.

Compound 2-T8 comprises abrin conjugated to SEQ ID NO:2.

Compound 2-T9 comprises abrin A chain (abrin toxin) conjugated to SEQ IDNO:2.

Compound 2-T10 comprises cobra venom factor (CVF) conjugated to SEQ IDNO:2.

Compound 2-T11 comprises gelonin (GEL) conjugated to SEQ ID NO:2.

Compound 2-T12 comprises saporin (SAP) conjugated to SEQ ID NO:2.

Compound 2-T13 comprises modeccin conjugated to SEQ ID NO:2.

Compound 2-T14 comprises viscumin conjugated to SEQ ID NO:2.

Compound 2-T15 comprises volkensin conjugated to SEQ ID NO:2.

Compounds 3-T1 to 3-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 3-T1 to 3-T15 each comprise SEQ IDNO:3 as the ST receptor binding moiety.

Compounds 5-T1 to 5-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 5-T1 to 5-T15 each comprise SEQ IDNO:5 as the ST receptor binding moiety.

Compounds 6-T1 to 6-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 6-T1 to 6-T15 each comprise SEQ IDNO:6 as the ST receptor binding moiety.

Compounds 7-T1 to 7-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 7-T1 to 7-T15 each comprise SEQ IDNO:7 as the ST receptor binding moiety.

Compounds 8-T1 to 8-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 8-T1 to 8-T15 each comprise SEQ IDNO:8 as the ST receptor binding moiety.

Compounds 9-T1 to 9-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 9-T1 to 9-T15 each comprise SEQ IDNO:9 as the ST receptor binding moiety.

Compounds 10-T1 to 10-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 10-T1 to 10-T15 each comprise SEQ IDNO:2 as the ST receptor binding moiety.

Compounds 11-T1 to 11-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 11-T1 to 11-T15 each comprise SEQ IDNO:11 as the ST receptor binding moiety.

Compounds 12-T1 to 12-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 12-T1 to 12-T15 each comprise SEQ IDNO:12 as the ST receptor binding moiety.

Compounds 13-T1 to 13-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 13-T1 to 13-T15 each comprise SEQ IDNO:13 as the ST receptor binding moiety.

Compounds 14-T1 to 14-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 14-T1 to 14-T15 each comprise SEQ IDNO:14 as the ST receptor binding moiety.

Compounds 15-T1 to 15-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 15-T1 to 15-T15 each comprise SEQ IDNO:15 as the ST receptor binding moiety.

Compounds 15-T1 to 15-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 15-T1 to 15-T15 each comprise SEQ IDNO:15 as the ST receptor binding moiety.

Compounds 17-T1 to 17-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 17-T1 to 17-T15 each comprise SEQ IDNO:17 as the ST receptor binding moiety.

Compounds 18-T1 to 18-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 18-T1 to 18-T15 each comprise SEQ IDNO:18 as the ST receptor binding moiety.

Compounds 19-T1 to 19-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 19-T1 to 19-T15 each comprise SEQ IDNO:19 as the ST receptor binding moiety.

Compounds 20-T1 to 20-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 20-T1 to 20-T15 each comprise SEQ IDNO:20 as the ST receptor binding moiety.

Compounds 21-T1 to 21-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 21-T1 to 21-T15 each comprise SEQ IDNO:21 as the ST receptor binding moiety.

Compounds 22-T1 to 22-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 22-T1 to 22-T15 each comprise SEQ IDNO:22 as the ST receptor binding moiety.

Compounds 23-T1 to 23-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 23-T1 to 23-T15 each comprise SEQ IDNO:23 as the ST receptor binding moiety.

Compounds 24-T1 to 24-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 24-T1 to 24-T15 each comprise SEQ IDNO:24 as the ST receptor binding moiety.

Compounds 25-T1 to 25-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 25-T1 to 25-T15 each comprise SEQ IDNO:25 as the ST receptor binding moiety.

Compounds 26-T1 to 26-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 26-T1 to 26-T15 each comprise SEQ IDNO:26 as the ST receptor binding moiety.

Compounds 27-T1 to 27-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 27-T1 to 27-T15 each comprise SEQ IDNO:27 as the ST receptor binding moiety.

Compounds 28-T1 to 28-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 28-T1 to 28-T15 each comprise SEQ IDNO:28 as the ST receptor binding moiety.

Compounds 29-T1 to 29-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 29-T1 to 29-T15 each comprise SEQ IDNO:29 as the ST receptor binding moiety.

Compounds 30-T1 to 30-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 30-T1 to 30-T15 each comprise SEQ IDNO:30 as the ST receptor binding moiety.

Compounds 31-T1 to 31-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 31-T1 to 31-T15 each comprise SEQ IDNO:31 as the ST receptor binding moiety.

Compounds 32-T1 to 32-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 32-T1 to 32-T15 each comprise SEQ IDNO:32 as the ST receptor binding moiety.

Compounds 33-T1 to 33-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 33-T1 to 33-T15 each comprise SEQ IDNO:33 as the ST receptor binding moiety.

Compounds 34-T1 to 34-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 34-T1 to 34-T15 each comprise SEQ IDNO:34 as the ST receptor binding moiety.

Compounds 35-T1 to 35-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 35-T1 to 35-T15 each comprise SEQ IDNO:35 as the ST receptor binding moiety.

Compounds 36-T1 to 36-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 36-T1 to 36-T15 each comprise SEQ IDNO:36 as the ST receptor binding moiety.

Compounds 37-T1 to 37-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 37-T1 to 37-T15 each comprise SEQ IDNO:37 as the ST receptor binding moiety.

Compounds 38-T1 to 38-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 38-T1 to 38-T15 each comprise SEQ IDNO:38 as the ST receptor binding moiety.

Compounds 39-T1 to 39-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 39-T1 to 39-T15 each comprise SEQ IDNO:39 as the ST receptor binding moiety.

Compounds 40-T1 to 40-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 40-T1 to 40-T15 each comprise SEQ IDNO:40 as the ST receptor binding moiety.

Compounds 41-T1 to 41-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 41-T1 to 41-T15 each comprise SEQ IDNO:41 as the ST receptor binding moiety.

Compounds 42-T1 to 42-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 42-T1 to 42-T15 each comprise SEQ IDNO:42 as the ST receptor binding moiety.

Compounds 43-T1 to 43-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 43-T1 to 43-T15 each comprise SEQ IDNO:43 as the ST receptor binding moiety.

Compounds 44-T1 to 44-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 44-T1 to 44-T15 each comprise SEQ IDNO:44 as the ST receptor binding moiety.

Compounds 45-T1 to 45-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 45-T1 to 45-T15 each comprise SEQ IDNO:45 as the ST receptor binding moiety.

Compounds 46-T1 to 46-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 46-T1 to 46-T15 each comprise SEQ IDNO:46 as the ST receptor binding moiety.

Compounds 47-T1 to 47-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 47-T1 to 47-T15 each comprise SEQ IDNO:47 as the ST receptor binding moiety.

Compounds 48-T1 to 48-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 48-T1 to 48-T15 each comprise SEQ IDNO:48 as the ST receptor binding moiety.

Compounds 49-T1 to 49-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 49-T1 to 49-T15 each comprise SEQ IDNO:49 as the ST receptor binding moiety.

Compounds 50-T1 to 50-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 50-T1 to 50-T15 each comprise SEQ IDNO:50 as the ST receptor binding moiety.

Compounds 51-T1 to 51-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 51-T1 to 51-T15 each comprise SEQ IDNO:51 as the ST receptor binding moiety.

Compounds 52-T1 to 52-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 52-T1 to 52-T15 each comprise SEQ IDNO:52 as the ST receptor binding moiety.

Compounds 53-T1 to 53-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 53-T1 to 53-T15 each comprise SEQ IDNO:53 as the ST receptor binding moiety.

Compounds 54-T1 to 54-T15 are the same as compounds 2-T1 to 2-T15,respectively, except instead of comprising SEQ ID NO:2 as the STreceptor binding moiety, compounds 54-T1 to 54-T15 each comprise SEQ IDNO:54 as the ST receptor binding moiety.

Compounds 2-AP, 3-AP and 5-AP to 54-AP refer to the 51 conjugatedcompounds that comprise alkaline phosphatase conjugated to SEQ ID NO:2,SEQ ID NO:3 and SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-NIZ, 3-NIZ and 5-NIZ to 54-NIZ refer to the 51 conjugatedcompounds that comprise nitroimidazole conjugated to SEQ ID NO:2, SEQ IDNO:3 and SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-MEZ, 3-MEZ and 5-MEZ to 54-MEZ refer to the 51 conjugatedcompounds that comprise metronidazole conjugated to SEQ ID NO:2, SEQ IDNO:3 and SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-MIS, 3-MIS and 5-MIS to 54-MIS refer to the 51 conjugatedcompounds that comprise misonidazole conjugated to SEQ ID NO:2, SEQ IDNO:3 and SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-47Sc, 3-47Sc and 5-47Sc to 54-47Sc refer to the 51conjugated compounds that comprise ⁴⁷Sc conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-67Cu, 3-67Cu and 5-67Cu to 54-67Cu refer to the 51conjugated compounds that comprise ⁶⁷Cu conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-90Y, 3-90Y and 5-90Y to 54-90Y refer to the 51 conjugatedcompounds that comprise ⁹⁰Y conjugated to SEQ ID NO:2, SEQ ID NO:3 ANDSEQ ID NO:5 through SEQ ID NO: 54, respectively.

Compounds 2-109Pd, 3-109Pd and 5-109Pd to 54-109Pd refer to the 51conjugated compounds that comprise ¹⁰⁹Pd conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-123I, 3-123I and 5-123I to 54-123I refer to the 51conjugated compounds that comprise ¹²³I conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-125I, 3-125I and 5-125I to 54-125I refer to the 51conjugated compounds that comprise ¹²⁵I conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-131I, 3-131I and 5-131I to 54-131I refer to the 51conjugated compounds that comprise ¹³¹I conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-132I, 3-132I and 5-132I to 54-132I refer to the 51conjugated compounds that comprise ¹³²I conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-186Re, 3-186Re and 5-186Re to 54-186Re refer to the 51conjugated compounds that comprise ¹⁸⁶Re, conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-188Re, 3-188Re and 5-188Re to 54-188Re refer to the 51conjugated compounds that comprise ¹⁸⁸Re, conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-199Au, 3-199Au and 5-199Au to 54-199Au refer to the 51conjugated compounds that comprise ¹⁹⁹Au, conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-211At, 3-211At and 5-211At to 54-211At refer to the 51conjugated compounds that comprise ²¹¹At, conjugated to SEQ ID NO:2, SEQID NO:3 AND, SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-212Pb, 3-212Pb and 5-212Pb to 54-212Pb refer to the 51conjugated compounds that comprise ²¹²Pb conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-212Bi, 3-212Bi and 5-212Bi to 54-212Bi refer to the 51conjugated compounds that comprise ²¹²Bi conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-203Pb, 3-203Pb and 5-203Pb to 54-203Pb refer to the 51conjugated compounds that comprise ²⁰³Pb conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-206Bi, 3-206Bi and 5-206Bi to 54-206Bi refer to the 51conjugated compounds that comprise ²⁰⁶Bi conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-32P, 3-32P and 5-32P to 54-32P refer to the 51 conjugatedcompounds that comprise ³²P conjugated to SEQ ID NO:2, SEQ ID NO:3 ANDSEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-33P, 3-33P and 5-33P to 54-33P refer to the 51 conjugatedcompounds that comprise ³³P conjugated to SEQ ID NO:2, SEQ ID NO:3 ANDSEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-71Ge, 3-71Ge and 5-71Ge to 54-71Ge refer to the 51conjugated compounds that comprise ⁷¹Ge conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-77As, 3-77As and 5-77As to 54-77As refer to the 51conjugated compounds that comprise ⁷⁷As conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-103Pd, 3-103Pd and 5-103Pd to 54-103Pd refer to the 51conjugated compounds that comprise ¹⁰³Pd conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-105Rh, 3-105Rh and 5-105Rh to 54-105Rh refer to the 51conjugated compounds that comprise ¹⁰⁵Rh conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-111Ag, 3-111Ag and 5-111Ag to 54-111Ag refer to the 51conjugated compounds that comprise ¹¹¹Ag conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-119Sb, 3-119Sb and 5-119Sb to 54-119Sb refer to the 51conjugated compounds that comprise ¹¹⁹Sb conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-121Sn, 3-121-Sn and 5-121Sn to 54-121Sn refer to the 51conjugated compounds that comprise ¹²¹Sn conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-131Cs, 3-131Cs and 5-131Cs to 54-131Cs refer to the 51conjugated compounds that comprise ¹³¹Cs conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-127Cs, 3-131Cs and 5-131Cs to 54-127Cs refer to the 51conjugated compounds that comprise ¹²⁷Cs conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-129Cs, 3-129Cs and 5-129Cs to 54-129Cs refer to the 51conjugated compounds that comprise ¹²⁹Cs conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-143Pr, 3-143Pr and 5-143Pr to 54-143Pr refer to the 51conjugated compounds that comprise ¹⁴³Pr conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-161Tb, 3-161Tb and 5-161Tb to 54-161Tb refer to the 51conjugated compounds that comprise ¹⁶¹Tb conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-177Lu, 3-177Lu and 5-177Lu to 54-177Lu refer to the 51conjugated compounds that comprise ¹⁷⁷Lu conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-191Os, 3-191Os and 5-191Os to 54-191Os refer to the 51conjugated compounds that comprise ¹⁹¹Os conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-193mPt, 3-193mPt and 5-193mPt to 54-193mPt refer to the 51conjugated compounds that comprise ^(193M)Pt conjugated to SEQ ID NO:2,SEQ ID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-197Hg, 3-197Hg and 5-197Hg to 54-197Hg refer to the 51conjugated compounds that comprise ¹⁹⁷Hg conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-43K, 3-43K and 5-43K to 54-43K refer to the 51 conjugatedcompounds that comprise ⁴³K conjugated to SEQ ID NO:2, SEQ ID NO:3 ANDSEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-52Fe, 3-52Fe and 5-52Fe to 54-52Fe refer to the 51conjugated compounds that comprise ⁵²Fe conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-57Co, 3-57Co and 5-57Co to 54-57Co refer to the 51conjugated compounds that comprise ⁵⁷Co conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-67Ga, 3-67Ga and 5-67Ga to 54-67Ga refer to the 51conjugated compounds that comprise ⁶⁷Ga conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-68Ga, 3-68Ga and 5-68Ga to 54-68Ga refer to the 51conjugated compounds that comprise ⁶⁸Ga conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-77Br, 3-77Br and 5-77Br to 54-77Br refer to the 51conjugated compounds that comprise ⁷⁷Br conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-81Rb, 3-81Rb and 5-81Rb to 54-81Rb refer to the 51conjugated compounds that comprise ⁸¹Rb conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-81mKr, 3-81mKr and 5-81mKr to 54-81mKr refer to the 51conjugated compounds that comprise ^(81M)Kr conjugated to SEQ ID NO:2,SEQ ID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-87mSr, 3-87mSr and 5-87mSr to 54-87mSr refer to the 51conjugated compounds that comprise ^(87M)Sr conjugated to SEQ ID NO:2,SEQ ID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-99mTc, 3-99mTc and 5-99mTc to 54-99mTc refer to the 51conjugated compounds that comprise ^(99M)Tc conjugated to SEQ ID NO:2,SEQ ID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-111In, 3-111In and 5-111In to 54-111In refer to the 51conjugated compounds that comprise ¹¹¹In conjugated to SEQ ID NO:2, SEQID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

Compounds 2-113mIn, 3-113mIn and 5-113mIn to 54-113mIn refer to the 51conjugated compounds that comprise ^(113M)In conjugated to SEQ ID NO:2,SEQ ID NO:3 AND SEQ ID NO:5 through SEQ ID NO:54, respectively.

The compounds described in this example are combined with apharmaceutically acceptable carrier or diluent to produce pharmaceuticalcompositions according to the present invention. Radiostable compoundsdescribed herein are useful in pharmaceutical compositions astherapeutics in the treatment of individuals suspected of suffering frommetastasized colorectal cancer including treatment of individualsdiagnosed with localized colorectal cancer as a prophylactic/therapeuticbefore metastasis can be readily detected. When present intherapeutically effective amounts, radioactive compounds describedherein are useful in pharmaceutical compositions as therapeutic agentsin the treatment of individuals suspected of suffering from metastasizedcolorectal cancer including treatment of individuals diagnosed withlocalized colorectal cancer as a prophylactic/therapeutic beforemetastasis can be readily detected. When present in diagnosticallyeffective amounts, radioactive compounds described herein are useful inpharmaceutical compositions as imaging agents in the diagnosis andidentification of metastasized colorectal cancer in individuals.

Example 2

One procedure for crosslinking ST receptor ligands which have a freeamino group such as ST receptor binding peptides, as for example SEQ IDNO:2, SEQ ID NO:3, and SEQ ID NOS:5-54 to active agents which have afree amino group such as methotrexate, doxorubicin, daunorubicin,cytosinarabinoside, cis-platin, vindesine, mitomycin and bleomycin, oralkaline phosphatase, or protein- or peptide-based toxin employshomobifunctional succinimidyl esters, preferably with chain carbonspacers such as disuccinimidyl suberate (Pierce Co, Rockford, Ill.).This approach of amino group derivatization has been employedsuccessfully to crosslink native ST to biotin and, ultimately, to largeagarose beads of micron-scale size, preserving the function of native ST(Hughes, M., et al. (1991) Biochem. 30:10738; Hakki, S., et al. (1993)Int. J. Biochem. 25:557; Almenoff, J. S., et al. (1992) Mol. Micro.8:865; each of which is incorporated herein by reference).

An ST binding ligand with the free amino group such as an ST receptorbinding peptide is incubated in the presence of the chemicalcrosslinking agent and an active agent which have a free amino group inequimolar quantities at room temperature for 15-30 min. Incubation isterminated by separating the reactants by gel permeation chromatographyby HPLC. This technique separates the conjugated compounds from freeactive agents and free ST binding ligands, active agent-active agentconjugates and ST binding ligand-ST binding ligand conjugates.Homogeneous preparations of conjugated through their free amino groupsand with a preferred molar ratio of 1:1 are obtained. As indicatedabove, complexing the free amino group of an ST peptide preservesreceptor binding function.

Example 3

In the event that a cleavable conjugated compound is required, the sameprotocol as described above may be employed utilizing3,3′-dithiobis(sulfosuccinimidylpropionate (SPDP); Pierce, IL.). SPDPforms a sulfhydral group from a free amino group which may be used toconjugate a compound to another free amino group. For example, STpeptides such as SEQ ID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 arederivatized using established procedures employing N-succinimidyl-3(2-pyridildithio)-propionate (SPDP, Pharmacia-LKB, NJ). The ST peptideis incubated with a 5-fold molar excess of SPDP for 30 minutes at roomtemperature. The ST-pyridylthiopropionate conjugate is separated fromunreacted reagents by gel permeation chromatography by HPLC. An activeagent with a free amino group, such as a protein-based toxin, isprepared for conjugation by reduction with dithiothreitol for 4 hours atroom temperature. Reduced active agent is incubated with a 2-fold molarexcess of ST receptor ligand-PDP conjugate at pH 8.0 for 36 hours at 4°C. Conjugate compound is purified from unreacted agents by gelpermeation chromatography by HPLC.

This protocol for conjugation is particularly useful to conjugate STpeptides to diphtheria toxin A chains and Pseudomonas exotoxin as wellas ricin toxin A chains (Magerstadt, M. Antibody Conjugates andMalignant Disease. (1991) CRC Press, Boca Raton, USA, pp. 110-152;Cawley, D. B. et al. (1980) Cell 22:563; Cumber, A. J., et al. (1985)Meth. Enz. 112:207; Gros, O. (1985) J. Immunol. Meth. 81:283; Worrell,N. R., et al. (1986) Anti-Cancer Drug Design 1: 179; Thorpe, P. E. etal. (1987) Cancer Res. 47:5924, each of which is incorporated herein byreference).

Example 4

Active agents with a free amino group may be derivatized with SPDP asdescribed above and conjugated with an ST ligand that has a free aminogroup and that has been modified with the succinimidyl ester ofiodoacetic acid (Pierce Co., Rockford, Ill.) (Magerstadt, M. (1991)Antibody Conjugates And Malignant Disease, CRC Press Boca Raton; Cumber,A. J. et al. (1985) Meth. Enz. 112:20, which are incorporated herein byreference). Conjugation relies on the selective reaction of iodoacetylgroups introduced into the amino terminal of the ST ligand with thethiol groups introduced into the active agent. As with the aboveprotocol, this procedure avoids homopolymer formation. However, theproduct is conjugated through a central thioether linkage which cannotbe reduced.

Example 5

An ST receptor ligand with a free amino group and active agents withfree amino groups may be conjugated through a disulfide bond usingiminothiolane (Pierce, Rockford, Ill.) (Fitzgerald, D. J. P. et al.(1983) Cell 32:607; Magerstadt, M. (1991) Antibody Conjugates AndMalignant Disease, CRC Press, Boca Raton; Bjorn, M. J., et al. (1985)Cancer Res. 45:1214; Bjorn, M. J., et al. (1986) Cancer Res. 46:3262,which are incorporated herein by reference). The ST receptor ligand witha free amino group is derivatized at the amino terminal withiminothiolane and the active agent is derivatized with SPDP as describedabove. Reacting iminothiolane-derivatized ST receptor ligand withSPDP-derivatized active agent results in conjugation by a reducibledisulfide bond. In addition, iminothiolane provides the versatility toconjugate these proteins through bonds other than disulfides. Thus,derivatization of active agents with the heterobifunctional agentsulfosuccinimidyl 4-(N-maleimidomethyl)cyclohexane (Pierce, Rockford,Ill.) and reaction with iminothiolane-derivatized ST receptor ligandwill conjugate these peptides without formation of disulfides.

Example 6

Conjugated compounds according to the invention which comprise an activemoiety that is a therapeutic agent specifically inhibit T84 cells invitro. The following protocols may be used to demonstrate that theconjugated compounds according to the invention which comprise an activemoiety that is chemotherapeutic or toxin specifically inhibit T84 cellsin vitro. Inhibition of T84 cells is assessed by determining the effectsof conjugated compounds on the ability of T84 cells to incorporate³⁵S-leucine into protein, ³H-thymidine into DNA, and to form colonies.The assessment of protein and DNA synthesis are classical techniques todetermine the cytotoxicity of conjugated compounds in vitro. Inhibitionof protein synthesis is measured because the toxins used as activemoieties are specific inhibitors of this process. Therefore, theseassays are the most sensitive measure of whether conjugated compoundsare binding to and internalized into T84 cells. Inhibition of DNAsynthesis is measured because some chemotherapeutics inhibit DNAsynthesis and further, it is a cytotoxicity assay which correlatesclosely with the reproductive survivability of cells in culture.Cytotoxicity, or the disruption of normal cellular metabolic processes,may not always directly correlate with cell survivability. Therefore,assessment of colony formation will directly measure the ability of theexperimental agents to decrease the survivability of tumor cells, whichclosely correlates with the impact of therapeutic agents on tumorviability in vivo. Controls include performing the same assay using theunconjugated form of the active agent and the unconjugated form of theST receptor ligand of which the conjugated compound is comprised inplace of the conjugated compound. The results obtained in the testassays and control assays are compared.

Conjugated compounds are assessed for their ability to inhibit proteinand DNA synthesis in vitro and to inhibit survival and proliferation bymeasuring colony formation in monolayer culture by established protocols(Wilson, A. P. (1987) “Cytotoxicity and viability assays”, Animal CellCulture: A Practical Approach. Freshney, R. I., ed. pp. 183-216, IRLPress, Oxford. which is incorporated herein by reference).

To assess the ability of a conjugated compound to inhibit proteinsynthesis in vitro, cells are plated in 200 μl of medium at asub-confluent density of 1-2×10⁵ and allowed to attach to form adividing cell monolayer over 12 hours at 37° C. Subsequently, the mediais replaced with 200 μl of fresh media containing the appropriateconcentration of conjugated compounds and cells incubated at 37° C. forvarious amounts of time. At the end of the indicated incubation period,cells is washed twice with medium and incubated at 37° C. in 0.5 ml ofmethionine-free medium supplemented with 0.5 μCi of L³⁵S-methionine (800Ci/mmol). After incubation for another 2 hours at 37° C., the medium isaspirated, cells washed twice with medium containing 1 mg/ml ofmethionine, and then precipitated in 12% ice-cold TCA. Radioactivityrecovered in TCA precipitates by centrifugation is quantified by liquidscintillation spectroscopy. In these studies, cells are maintained inlog growth and assays are performed using triplicate wells. Data isexpressed as a percentage of protein synthesis observed in the presenceof experimental agents compared to untreated cells.

To assess the ability of a conjugated compound to inhibit DNA synthesisin vitro cells are plated as a subconfluent monolayer and incubated withexperimental agents as described above. At the end of the incubationperiod, cells are washed twice and incubated at 37° C. in mediumcontaining 2.5 μCi of ³H-thymidine (5 Ci/mmol). After incubation foranother hour, cells are processed with TCA, precipitates recovered, andradioactivity quantified as described above. As above, cells aremaintained in log growth and assays is performed in triplicate. Data isexpressed as a percentage of DNA synthesis observed in the presence ofexperimental agents compared to untreated cells.

To assess the ability of a conjugated compound to inhibit survival andproliferation by measuring colony formation in monolayer culture, cellsare plated as a sub-confluent monolayer on 25 cm² flasks and allowed toattach as described above. The medium is replaced with that containingvarious concentrations of experimental agents and incubated with cellsfor various amounts of time. At the end of the incubation, cells arerecovered as a single cell suspension by trypsinization and replated toa density which will yield 100-200 colonies per 6 cm plate. Cells arepermitted to grow for 7 days, then fixed in methanol, stained with 1%crystal violet, and the number of colonies quantified. Assays areperformed in duplicate and data is expressed as a percentage of colonyformation observed in the presence of experimental agents compared tountreated cells. Results in our laboratory have demonstrated that T84cells can be placed into single cell suspensions utilizing trypsin (10μg/ml) with a plating efficiency of 40% and a doubling time of 18 hours.

Example 7

Radioactive iodine such as ¹²³I, ¹²⁵I, ¹³¹I and ¹³²I, can be added to anST receptor binding peptide such as an ST peptide using a standardprotocol well-known to those having ordinary skill in the art (Thompson,M. et al. (1985) Analytical Biochemistry 148:26, which is incorporatedherein by reference). Radioactive iodine is conjugated directly to an STpeptide such as SEQ ID NO:2, SEQ ID NO:3 or SEQ ID NO:5 at tyrosine-5,tyrosine-4 or tyrosine-5, respectively.

Briefly, the ST peptide is produced in bacteria. For example, E. colistrain 431 is grown in culture and secretes ST into this culture. Theculture media is then purified using routine techniques. ST can also bemade by solid-phase synthesis as has been done previously, usingstandard techniques. (Dreyfus, L., et al. (1983) Infec. Immun. 42:539,which is incorporated herein by reference.

Ten micrograms of ST peptide are reacted with 2 milliCuries ofradioactive INa (Amersham Corporation, Massachusetts) in the presence ofIodobeads (Bio Rad Laboratories, CA) and beta-D-glucose. These arereacted for 30 min after which the products are subjected tochromatography on a Sepak reversed-phase cartridge (Millipore Corp., MA)followed by separation on a C₁₈reversed-phase column by HPLC using a20-25% acetonitrile gradient. Conjugated compositions which comprise SEQID NO:2, SEQ ID NO:3 or SEQ ID NO:5 with the radioiodine attached totyrosine-4 elutes at 45 min. These molecules retain full biochemical andpharmacological activity.

Example 8

¹²⁵I is conjugated directly to an ST peptide such as SEQ ID NO:13 attyrosine-4.

SEQ ID NO:13 is produced by solid-phase synthesis as described above.Ten micrograms of SEQ ID NO:13 are reacted with 2 milliCuries of ¹²⁵INa(Amersham Corporation, Massachusetts) in the presence of Iodobeads (BioRad Laboratories, CA) and beta-D-glucose. These are reacted for 30 minafter which the products are subjected to chromatography on a Sepakreversed-phase cartridge (Millipore Corp., MA) followed by separation ona C₁₈reversed-phase column by HPLC using a 20-25% acetonitrile gradient.¹²⁵I-SEQ ID NO:13 conjugate with the radioiodine attached to tyrosine-4elutes at 45 min. This molecule retains full biochemical andpharmacological activity.

Dosing of radioiodine for diagnostic imaging typically requires about 4milliCuries/patient (Steinstraber, A., et al. (1988) J. Nucl. Med.29:875; Wessels, B. W. and Rogus, R. D. (1984) Med. Phys. 11:638; Kwok,C. S., et al. (1985) Med. Phys. 12:405). For proteins labeled with aspecific activity of 2,000 Curies/mmol, such as ST peptide, this wouldrequire about 10 micrograms of labeled peptide injected intravenouslyper patient for diagnostic imaging.

Example 9

¹³¹I is conjugated directly to an ST peptide such as SEQ ID NO:13 attyrosine-4.

SEQ ID NO:13 is produced by solid-phase synthesis as described above.Ten micrograms of SEQ ID NO:13 are reacted with 10 milliCuries of ¹³¹INa(Amersham Corporation, Massachusetts) in the presence of Iodobeads (BioRad Laboratories, CA) and beta-D-glucose. These are reacted for 30 minafter which the products are subjected to chromatography on a Sepakreversed-phase cartridge (Millipore Corp., MA) followed by separation ona C₁₈reversed-phase column by HPLC using a 20-25% acetonitrile gradient.¹³¹I-SEQ ID NO:13 conjugate with the radioiodine attached to tyrosine-4elutes at 45 min. This molecule retains full biochemical andpharmacological activity.

Typically, for radioiodinated antibodies (MW=160,000 Da), about 150nanomoles of protein (24 milligrams) labeled with a specific activity of10,000 Curies/mmol are required per gram of tumor per patient (Humm, J.L. (1986) J. Nucl. Med. 27:1490). Thus, for proteins labeled with aspecific activity of 2,000 Curies/mmol, with a molecular weight of 2,000Da, such as ST peptide, about 3 milligrams would be required per gram oftumor per patient for intravenous infusion.

Example 10

In some embodiments, coupling of ST receptor ligands which have a freeamino group, particularly ST receptor binding peptides such as STpeptides, and active agents with a free amino group such asprotein-based toxins is performed by introducing a disulfide bridgebetween the 2 molecules. This strategy is particularly useful toconjugate ST peptides since the free amino terminal has been shown to beuseful as a point of conjugation without affecting ST binding activity.This strategy is particularly useful to conjugate protein-based toxinssince the free amino terminal is available on such molecules and forsome conjugated compounds, most notably RTA conjugates, a disulfidebride which can be reduced to yield separate proteins has beendemonstrated to be important in the construction of functional chimerastargeted by monoclonal antibodies (Magerstadt, M. (1991) AntibodyConjugates And Malignant Disease, CRC Press, Boca Raton; Bjorn, M. J.,et al. (1985) Cancer Res. 45:1214; Bjorn, M. J., et al. (1986) CancerRes. 46:3262; Masuho, Y., et al. (1982) J. Biochem. 91:1583, which areeach incorporated herein by reference). While some toxins may be coupledto ST peptides using crosslinking agents which do not result in areducible disulfide bridge between the individual components but retainfunctional cytotoxicity, ricin A chain toxin requires a reducibledisulfide for cytotoxicity while Pseudomonas exotoxin, for example, doesnot.

Disulfide coupling is achieved using established procedures employingthe heterobifunctional agent N-succinimidyl-3(2-pyridyldithio)-proportionate (SPDP, Pharmacia-LKB, Piscataway, N.J.)(Magerstadt, M. (1991) Antibody Conjugates And Malignant Disease, CRCPress, Boca Raton; Cawley, D. B. et al. (1980) Cell 22:563; Cumber, A.J., et al. (1985) Meth. Enz. 112:20; Gros, O., et al. (1985) J. Immunol.Meth. 81:283; Worrell, N. R., (1986) Anti-Cancer Drug Design 1:19;Thorpe, P. E., et al. (1987) Cancer Res. 4:5924, which are incorporatedherein by reference).

In some embodiments, toxins including the A chains of deglycosylatedricin toxin (RTA; Sigma Chemical Co., St. Louis, Mo.), diphtheria toxinA (DTA; Calbiochem, La Jolla, Calif.) and Pseudomonas exotoxin (PEA) areconjugated to ST peptides to produce conjugated compositions accordingto the present invention using this procedure. Deglycosylated RTA isemployed since the glycosylated form of this toxin exhibits non-specificbinding to liver cells. DTA is prepared from diphtheria toxin by anestablished procedure (Michel, A. and Drykx, J. (1975) Biochem. Biophys.Acta 365:15; Cumber, A. J., et al. (1985) Meth. Enz. 112:207, both ofwhich are incorporated herein by reference). PEA

In some embodiments, ST peptides are conjugated to toxins by thisprocedure. For example, the ST peptide SEQ ID NO:3 which is produced asdescribed above (see Dreyfus, L., et al. (1983) Infec. Immun. 42:539,which is incorporated herein by reference).

Toxins are prepared for coupling by reduction with 0.1 M dithiothreitol(DTT) for 4 hours at room temperature in 0.4 M Tris-HCl, pH 8.0 and 1 mMEDTA. Reduced toxins are desalted on a Sephadex G-25 column equilibratedin TES buffer and mixed with a 2-fold molar excess of ST-PDP. Reactionsare adjusted to pH 8.0 with TES and incubated at 4° C. for 36 hours. STpeptide-toxin conjugates are purified from unreacted products andhomopolymers of ST peptides and toxins by gel filtration on SephadexG-75 in 20 mM TES, pH 8.0 containing 0.1 M NaCl. Chromatographicfractions are monitored by SDS-PAGE on 10% polyacrylamide gels undernon-reducing conditions for the presence of 1:1 conjugates of STpeptides and toxins. Also, these conjugates are analyzed by 10% SDS-PAGEunder reducing conditions, to insure that ST and cytotoxins are coupledby a reducible disulfide bond. Molar concentrations of the conjugate arecalculated by quantifying radioactivity in these samples.

ST trace labeled with ¹²⁵I on tyrosine 4 (10 Ci/mmol) is used in orderto follow the conjugate through various separation and chromatographicsteps and to enable us to calculate the molar ratio of ST to cytotoxinin the final purified conjugate. ST trace labeled with 125I isderivatized by incubating 1 mg/ml with a 5-fold molar excess of SPDP for30 min at room temperature in Na phosphate buffer, pH 7.4. TheST-pyridylthiopropionate (ST-PDP) conjugate is purified from unreactedcrosslinking agent by chromatography on Sephadex G-25 equilibrated with20 mM N-Tris(hydroxymethyl)-methyl-2-aminoethane sulfonic acid (TES)buffer, pH 7.4. Preservation of receptor binding of conjugated STpeptides in human intestinal membranes is determined in competitionassays of increasing concentrations of ST-PDP and ¹²⁵I-ST (5×10¹⁰M), toinsure that this process does not destroy the function of the STreceptor ligand.

The above coupling protocol has several advantages for conjugating thevarious toxins. First, it introduces a reducible disulfide bridge intothe conjugated composition, important for RTA cytotoxicity. Also, thistechnique avoids the exposure of ST peptide to quantitative reductionwith DTT which could interrupt its 3 intrachain disulfide bondsimportant for receptor binding activity. In addition, there is a singlegroup available at the amino terminal of ST peptide for derivatizationwith SPDP and previous experiments have demonstrated that derivatizationof that group preserves the binding properties of the ligand. Therefore,other configurations for conjugation which could result in inactivationof ST are not possible. Furthermore, PEA requires preactivation with DTTto achieve optimum cytotoxicity which will be accomplished utilizing theabove protocol.

To produce a functional conjugated compound that comprises a toxin, itis essential that the receptor binding and enzyme activities of themoieties are preserved throughout the process of conjugation. Therefore,once such conjugate compounds are obtained, they are tested for thepreservation of those functions. ST receptor binding activity ofconjugated compounds is examined in competitive binding assays, asdescribed above. In these studies, increasing concentrations of theconjugated compounds are incubated with a constant concentration(5×10¹⁰M) of ¹²⁵I-ST and intestinal membranes (50-100 μg of protein) toachieve equilibrium. Parallel incubations contain excess (5×10⁷M)unlabeled ST to assess non-specific binding. The concentration-dependentcompetitive displacement of radiolabeled ST by conjugated compounds iscompared to the competitive displacement achieved by native ST.Displacement curves are employed to estimate the affinity of eachconjugated compound (K_(D)) and compare that to the affinity of nativeST measured by this technique. Control studies include evaluating theability of unconjugated toxins to compete with native ST for receptorbinding. These studies establish that the binding function of ST in theconjugated construct is preserved.

Preservation of toxin activity in conjugated compounds is also assessed.PEA and DTA induce toxicity by catalyzing the NAD-dependentADP-ribosylation of elongation factor 2 (EF2), inhibiting proteinsynthesis. ADP-ribosyl transferase activity is assessed using anestablished assay (Chung, D. W. and Collier, R. J. Infect. Immun.16:832; Fitzgerald, D. J. P. (1987) Meth. Enz. 151:139, which are bothincorporated herein by reference). Reactions are conducted in 30 MmTris-HCl, pH 8.2 containing 40 mM DTT, 50 mCi ¹⁴C-NAD, and 20 μl ofrabbit reticulocyte lysate containing elongation factor 2 (EF-2;Promega, Madison, Wis.) in a total volume of 500 μl. Reactions areinitiated by the addition of lysate, incubated for 30 minutes at 37° C.,and terminated by the addition of ice-cold 12% TCA. Radioactivity inprotein precipitates collected by centrifugation is quantified by liquidscintillation spectroscopy. The ability of the conjugated compounds thatcomprise DTA or PEA to catalyze the transfer of labeled ADP-ribose toEF-2 is compared to that catalyzed by similar quantities of unconjugatedtoxins. Control experiments include examining the ability ofunconjugated toxins or ST to catalyze ADP-ribose transfer and theeffects of ST on the enzymatic activity of unconjugated cytotoxins.

RTA inhibits protein synthesis by catalytically inactivating the 60Sribosomal subunit. The catalytic activity of conjugated compounds thatcomprise RTA is assessed by its ability to inhibit protein synthesis incell-free assays using established procedures (Leonard, J. E. et al.(1985) Cancer Res. 45:5263 which is incorporated herein by reference).Assays contain 35 μl of nuclease-treated rabbit reticulocyte lysates, 1μl of 1 mM mixed amino acids deficient in methionine, 2 μl of Bromemosaic RNA (Promega, Madison, Wis.) at 0.5 μg/μl, 7 μl of sterile wateror conjugate solution, and 5 μCi of ³⁵S-methionine in a total volume of50 μl. Reactions will be initiated by the addition of lysate, incubatedat 30° C. for 30 minutes, and terminated by the use of addition of 12%TCA. Radioactivity in protein precipitates collected by centrifugationis quantified by liquid scintillation spectroscopy. Control experimentsinclude examining the ability of unconjugated RTA or ST peptide toinhibit cell-free protein synthesis and the effects of ST peptide on theinhibitory activity of the unconjugated cytotoxin.

Example 11

Methotrexate is linked to SEQ ID NO:12 by the homobifunctionalcrosslinker succinimidyl esters with long chain carbon spacers such asdisuccinimidyl suberate (Pierce, IL). SEQ ID NO:12 is incubated in thepresence of the chemical crosslinking agent and methotrexate inequimolar quantities at room temperature for 15-30 min. Incubation isterminated by separating the reactants by gel permeation chromatographyby HPLC. This technique separates the methotrexate/SEQ ID NO:12conjugates from free drug, free ST peptide, drug-drug conjugates and STpeptide-ST peptide conjugates. Homogeneous preparations of SEQ IDNO:12-methotrexate conjugates coupled through their free amino groupsand with a preferred molar ratio of 1:1 are obtained. Complexing thefree amino group of ST preserves receptor binding function.

Example 12

¹¹¹In is coupled to SEQ ID NO:37 with functional amino groups using achelator. The ST peptide has a free amino function at the amino terminalwhich may be modified without altering the ST receptor binding activityof the ST peptide. ¹¹¹In is rapidly and potently chelated by either EDTA(ethylenediaminetetraacetic acid) or DTPA (diethylenetriaminepetaaceticacid). DTPA is preferred over EDTA because the latter may be moreunstable in vivo. The ¹¹¹In-DTPA is converted to a mixedN-hydroxysuccinimide ester which is reactive with free amino groups,mixed with ST, and the reaction products, including ¹¹¹In-SEQ ID NO:37separated by HPLC (Bremer, K. H. and Schwarz, A. (1987) in Safety AndEfficacy Of Radiopharmaceuticals. Kristensen, K. and Norbygaard, E.,Eds. Martinius Nijhoff, Dordrecht, The Netherlands, P. 43; Krejcarek, G.E., and Tucker, K. L. (1977) Biochem. Biophys. Res. Commun. 77:581;Paxton, R. J., et al. (1985) Cancer Res. 45:5694; Richardson, A. P., etal. (1986) Nucl. Med. Biol. 14:569, which are each incorporated hereinby reference).

Example 13

^(99m)Tc can be conjugated to SEQ ID NO:46 using an approach which issimilar to that for indium. Thus, technetium can be chelated by DTPAwhich is converted to an anhydride, such as N-hydroxysuccinimideanhydride, and reacted with SEQ ID NO:46. The ST-technetium conjugatecan then be separated using HPLC (Magerstadt, M. (1991) AntibodyConjugates And Malignant DiseaseCRC Press, Boca Raton; Eckelman, W. C.and Paik, C. H. (1986) Nucl. Med. Biol. 14:569)

Example 14

Diphtheria toxin A chain (DTA) is prepared from native diphtheria toxinby standard techniques. SEQ ID NO:22 is coupled to N-succinimidyl-3(2-pyridyldithio)-propionate (SPDP, Pharmacia-LKB, Piscataway, N.J.) andthe SEQ ID NO:22-PDP conjugate is purified by HPLC by establishedprocedures. DTA is reduced with dithiothreitol and incubated with SEQ IDNO:22-PDP. DTA-SEQ ID NO:22 is purified after conjugation using HPLC.

Example 15

Pseudomonas Exotoxin is prepared from native sources by standardtechniques. SEQ ID NO:54 is coupled toN-succinimidyl-3(2-pyridyldithio)-propionate (SPDP, Pharmacia-LKB,Piscataway, N.J.) and the SEQ ID NO:54-PDP conjugate is purified by HPLCby established procedures. Pseudomonas Exotoxin is reduced withdithiothreitol and incubated with SEQ ID NO:54-PDP. PseudomonasExotoxin-SEQ ID NO:54 is purified after conjugation using HPLC.

Example 16

Doxorubicin is linked to SEQ ID NO:54 by the homobifunctionalcrosslinker succinimidyl esters with long chain carbon spacers such asdisuccinimidyl suberate (Pierce, IL). SEQ ID NO:54 is incubated in thepresence of the chemical crosslinking agent and doxorubicin in equimolarquantities at room temperature for 15-30 min. Incubation is terminatedby separating the reactants by gel permeation chromatography by HPLC.This technique separates the doxorubicin/SEQ ID NO:54 conjugates fromfree doxorubicin, free ST peptide, drug-drug conjugates and STpeptide-ST peptide conjugates. Homogeneous preparations of SEQ IDNO:54-doxorubicin conjugates coupled through their free amino groups andwith a preferred molar ratio of 1:1 are obtained. Complexing the freeamino group of ST preserves receptor binding function.

Example 17

Daunorubicin is linked to SEQ ID NO:32 by the homobifunctionalcrosslinker succinimidyl esters with long chain carbon spacers such asdisuccinimidyl suberate (Pierce, IL). SEQ ID NO:32 is incubated in thepresence of the chemical crosslinking agent and daunorubicin inequimolar quantities at room temperature for 15-30 min. Incubation isterminated by separating the reactants by gel permeation chromatographyby HPLC. This technique separates the daunorubicin/SEQ ID NO:54conjugates from free daunorubicin, free ST peptide, drug-drug conjugatesand ST peptide-ST peptide conjugates. Homogeneous preparations of SEQ IDNO:54-daunorubicin conjugates coupled through their free amino groupsand with a preferred molar ratio of 1:1 are obtained. Complexing thefree amino group of ST preserves receptor binding function.

1. A conjugated compound comprising: a) a ST receptor binding moiety;and b) an active moiety; wherein said active moiety is a radiostableactive agent.
 2. The compound of claim 1 wherein said ST receptorbinding moiety is a peptide.
 3. The compound of claim 1 wherein said STreceptor binding moiety is selected from the group consisting of: SEQ IDNO:2, SEQ ID NO:3, SEQ ID NOS:5-54 and fragments and derivativesthereof.
 4. The compound of claim 1 wherein said ST receptor bindingmoiety is selected from the group consisting of SEQ ID:2, SEQ ID NO:3,SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:54.
 5. The compound of claim 1wherein said active moiety is a therapeutic agent.
 6. The compound ofclaim 1 wherein said active moiety is selected from the group consistingof: methotrexate, doxorubicin, daunorubicin, cytosinarabinoside,etoposide, 5-4 fluorouracil, melphalan, chlorambucil, cis-platin,vindesine, mitomycin, bleomycin, purothionin, macromomycin,1,4-benzoquinone derivatives, trenimon, ricin, ricin A chain,Pseudomonas exotoxin, diphtheria toxin, Clostridium perfringensphospholipases C, bovine pancreatic ribonuclease, pokeweek antiviralprotein, abrin, abrin A chain, cobra venom factor, gelonin, saporin,modeccin, viscumin, volkensin, alkaline phosphatase, nitroimidazole,metronidazole and misonidazole.
 7. The compound of claim 1 wherein: a)said ST receptor binding moiety is selected from the group consistingof: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NOS:5-54 and fragments andderivatives thereof; b) said an active moiety is selected from the groupconsisting of: methotrexate, doxorubicin, daunorubicin,cytosinarabinoside, etoposide, 5-4 fluorouracil, melphalan,chlorambucil, cis-platin, vindesine, mitomycin, bleomycin, purothionin,macromomycin, 1,4-benzoquinone derivatives, trenimon, ricin, ricin Achain, Pseudomonas exotoxin, diphtheria toxin, Clostridium perfringensphospholipases C, bovine pancreatic ribonuclease, pokeweed antiviralprotein, abrin, abrin A chain, cobra venom factor, gelonin, saporin,modeccin, viscumin, volkensin, alkaline phosphatase, nitroimidazole,metronidazole and misonidazole.
 8. The compound of claim 1 wherein saidan active moiety is selected from the group consisting of: methotrexate,doxorubicin, daunorubicin, cytosinarabinoside, cis-platin, vindesine,mitomycin and bleomycin, alkaline phosphatase, ricin A chain,Pseudomonas exotoxin and diphtheria toxin.
 9. The compound of claim 1wherein: a) said ST receptor binding moiety is selected from the groupconsisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:6 andSEQ ID NO:54; and b) said an active moiety is selected from the groupconsisting of: methotrexate, doxorubicin, daunorubicin,cytosinarabinoside, cis-platin, vindesine, mitomycin and bleomycin,alkaline phosphatase, ricin A chain, Pseudomonas exotoxin and diphtheriatoxin.
 10. A pharmaceutical composition comprising: a) apharmaceutically acceptable carrier or diluent, and b) a conjugatecompound according to claim
 1. 11. A method of treating an individualsuspected of suffering from metastasized colorectal cancer comprisingthe steps of administering to said individual a pharmaceuticalcomposition according to claim
 10. 12. A pharmaceutical compositioncomprising: a) a pharmaceutically acceptable carrier or diluent, and, b)conjugated compound comprising: i) a ST receptor binding moiety; and ii)an active moiety wherein said active moiety is a radioactive agent andsaid conjugated compound is present in an amount effective fortherapeutic or diagnostic use in humans suffering from colorectalcancer.
 13. The pharmaceutical composition of claim 12 wherein saidactive moiety is selected from the group consisting of: ⁴⁷Sc, ⁶⁷Cu, ⁹⁰Y,¹⁰⁹Pd, ¹²³I, ¹²⁵I, ¹³¹I, ¹⁸⁶Re, ¹⁹⁹Au, ²¹¹At, ²¹²Pb, ²¹²B, ³²P, ³³P,⁷⁷Ge, ⁷⁷As, ¹⁰³Pb, ¹⁰⁵Rh, ¹¹¹Ag, ¹¹⁹Sb, ¹²¹Sn, ¹³¹Cs, ¹⁴³Pr, ¹⁶¹Tb,¹⁷⁷Lu, ¹⁹¹Os, ^(193M)Pt and ¹⁹⁷Hg.
 14. The pharmaceutical composition ofclaim 12 wherein said active moiety is selected from the groupconsisting of: ⁴³K, ⁵²Fe, ⁵⁷Co, ⁶⁷Cu, ⁶⁷Ga, ⁶⁸Ga, ⁷⁷Br, ⁸¹Rb/^(81M)Kr,^(87M)Sr, ^(99M)Tc, ¹¹¹In, ¹¹³In, ¹²³I, ¹²⁵I, ¹²⁷Cs, ¹²⁹Cs, ¹³¹I, ¹³²I,¹⁹⁷Hg, ²⁰³Pb and ²⁰⁶Bi.
 15. The pharmaceutical composition of claim 12wherein said ST receptor binding moiety is a peptide.
 16. Thepharmaceutical composition of claim 12 wherein said ST receptor bindingmoiety is selected from the group consisting of: SEQ ID NO:2, SEQ IDNO:3, SEQ ID NOS:5-54 and fragments and derivatives thereof.
 17. Thepharmaceutical composition of claim 12 wherein said ST receptor bindingmoiety is selected from the group consisting of SEQ ID NO:2, SEQ IDNO:3, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:54.
 18. The pharmaceuticalcomposition of claim 12 wherein said ST receptor binding moiety isselected from the group consisting of: SEQ ID NO:2, SEQ ID NO:3, SEQ IDNO:5, SEQ ID NO:6 and SEQ ID NO:54, and said active moiety is selectedfrom the group consisting of: ⁴⁷Sc, ⁶⁷Cu, ⁹⁰Y, ¹⁰⁹Pd, ¹²³I, ¹²⁵I, ¹³¹I,¹⁸⁶Re, ¹⁸⁸Re, ¹⁹⁹Au, ²¹¹At, ²¹²Pb, ²¹²B, ³²P and ³³P, ⁷¹Ge, ⁷⁷As, ¹⁰³Pb,¹⁰⁵Rh, ¹¹¹Ag, ¹¹⁹Sb, ¹²¹Sn, ¹³¹Cs, ¹⁴³Pr, ¹⁶¹Tb, ¹⁷⁷Lu, ¹⁹¹Os, ^(193M)Ptand ¹⁹⁷Hg.
 19. The pharmaceutical composition of claim 12 wherein saidST receptor binding moiety is selected from the group consisting of: SEQID NO:2, SEQ ID NO:3, SEQ ID NO.5, SEQ ID NO:6 and SEQ ID NO:54, andsaid active moiety is selected from the group consisting of ⁴³K, ⁵³Fe,⁵⁷Co, ⁶⁷Cu, ⁶⁷Ga, ⁶⁸Ga, ⁷⁷Br, ⁸¹Rb/^(81M)Kr, ^(87M)Sr, ^(99M)Tc, ¹¹¹In,¹²³I, ¹²⁵I, ¹²⁷Cs, ¹²⁹Cs, ¹³¹I, ¹³²I, ¹⁹⁷Hg, ²⁰³Pb and ²⁰⁶Bi.
 20. Amethod of radioimaging metastasized colorectal cancer cells comprisingthe steps of administering to an individual a pharmaceutical compositioncomprising: a) a pharmaceutically acceptable carrier or diluent, and b)conjugated compound comprising: i) a ST receptor binding moiety; and ii)an active moiety; wherein said active moiety is a radioactive agent andsaid conjugated compound is present in an amount effective fordiagnostic use in humans suffering from colorectal cancer.
 21. A methodof treating an individual suspected of suffering from metastasizedcolorectal cancer comprising the steps of administering to saidindividual a pharmaceutical composition comprising: a) apharmaceutically acceptable carrier or diluent, and, b) conjugatedcompound comprising: i) a ST receptor binding moiety; and ii) an activemoiety; wherein said active moiety is a radiostable agent or radioactiveagent and said conjugated compound is present in an amount effective fortherapeutic or diagnostic use in humans suffering from colorectalcancer.
 22. A method of delivery of a nucleic acid molecule tointestinal tract cells of an individual comprising the steps ofadministering to said individual a pharmaceutical compositioncomprising: a) a pharmaceutically acceptable carrier or diluent, and, b)a composition comprising: i) a ST receptor ligand; and ii) a nucleicacid molecule.
 23. A method of treating an individual suspected ofsuffering from metastasized colorectal cancer comprising the steps ofadministering to said individual: a) a conjugated compound comprising anST receptor binding moiety and an enzyme; and b) a prodrug which whenprocessed by the enzyme is converted to a drug.
 24. The method of claims23 wherein: a) a conjugated compound comprises an ST receptor bindingmoiety and alkaline phosphatase; and b) the prodrug isetoposidephosphate.
 25. The compound of claim 1 wherein said activemoiety is an enzyme.
 26. The compound of claim 25 wherein said activemoiety is alkaline phosphatase.